2^nd Glycobiology World Congress August 29-31,2016 Atlanta, USA

Biography:

Dr. Razi is the founder of AccuDava Inc., a biomedical company for personalized cancer chemotherapy (http://www.accudava.com), based on the discovery of two Glycomarkers. Razi received her PhD in Medical Sciences from Uppsala University-Sweden, in 1995. In her post-doctoral program, at the University of California-San Diego, she discovered a novel glycan-mediated lymphocytes activation mechanism in the immune system. She then joined the Consortium for Functional Glycomics (CFG), at The-Scripps-Research-Institute, where her team successfully developed and launched the world’s largest mammalian glycan microarray in 2011. This unique platform that displays 611 defined glycans has since become available globally for studying glycan-mediated interactions.

Abstract:

Personalized chemotherapy is an unmet need in cancer treatments. A molecular test that can accurately predict the drug efficacy on a patient would be invaluable in selecting effective treatment strategy. We have identified two glycan structures, namely Glycomarker-1 and Glycomarker-2, whose expression levels on the cancer cell surface are associated with responses to chemotherapy with platinum drugs (US patent #7585503, and International pending patent). This finding introduces a new concept, linking glycan cell surface expressions with drug reactivity, and proposes a glycan-mediated mechanism for drug uptake. Our initial studies profiling the cell surface glycans, using Flow Cytometry with specific glycan-binding lectins, on three isogenic pairs of ovarian carcinoma cell-lines, consisting of chemosensitive and acquired chemoresistant phenotypes, revealed at least a ten-fold decrease in alpha2-6Sialyl-R motif (Glycomarker-1) on resistant phenotypes compared to sensitive cells. Further studies by fluorescent confocal microscopy, colony forming assay, sialidase treatments, and mass-spectrometry confirmed the association of Glycomarker-1 with drug uptake. Using Lectin histochemistry (LHC) on clinical samples proved a feasible assay for Glycomarker-1, tested on 64 human ovarian normal and cancerous tissue sections. The LHC on retrospective ovarian cancer specimens, with a known history of drug-response, correctly predicted drug-responses in 22 out of 27 (81.4%) patients. During the studies on the mechanism for drug response, another glycan structure, Glycomarker 2, was identified demonstrating a similar expression pattern to Glycomarker 1. Further studies on Glycomarker-2 suggest an association of the two glycomarkers that would put forward a glycan-mediated mechanism for platinum-drug uptake by cancer cells.

Biography:

For the past 35 years, Dr. Szewczuk is Full Professor of Immunology and Medicine, Queen’s University, Kingston, Ontario Canada. He received his B.Sc. in Chemistry (U. of Guelph), M.Sc. in Biochemistry (Guelph), Ph.D. in Immunochemistry (U. of Windsor) and post-doctoral training with Gregory W. Siskind, M.D. in cellular immunology at Cornell University Medical College, NYC. Dr. Szewczuk’s recent research has focused on the role of glycosylation in receptor activation with a particular focus of TOLL-like, nerve growth factor Trk, EGFR and insulin receptors. He has discovered a novel receptor-signaling platform and its targeted translation in multistage of tumorigenesis.

Abstract:

Multicellular tumor spheroids are now at the forefront of cancer research, designed to mimic tumor-like developmental patterns in vitro. Tumor growth in vivo is known to be highly influenced by aberrant cell surface specific sialoglycan structures on glycoproteins. Aberrant sialoglycan patterns that facilitate spheroid formation are not well defined. Here, matrix-free spheroids from human breast MCF-7 and pancreatic PANC1 cancer cell lines and their respective tamoxifen (TMX) and gemcitabine (Gem) resistant variants were generated using the RGD induced self-assembly platform. Pretreatment with specific lectins, exogenous neuraminidase dose dependently reduced spheroid volume formation. Oseltamivir phosphate (OP) promoted cell aggregation and compaction into spheroid formation. PANC1 and triple negative breast MDA-MB231 xenograft tumors from untreated and OP-treated RAGxCγ double mutant mice expressed significant higher levels of α-2,3-sialoglycan (SA) than α-2,6-SA. MCF-7 spheroids expressed higher α-2,3-SA to α-2,6-SA ratio. The relative levels of specific sialoglycan structures on the cell surface facilitate avascular 3D multicellular tumor spheroids in vitro as well as in vivo tumors.

Biography:

Iva Turyan completed her PhD at the age of 27 years from St. Petersburg University and postdoctoral studies at The Hebrew University of Jerusalem. She is currently an Analytical Development Scientist at Biogen, Cambridge, MA. She has published more than 45 papers in reputed journals, and has been awarded 6 patents.

Abstract:

Glycosylation of therapeutic recombinant proteins is of particular importance due to its potential impact on solubility, bioactivity, pharmacokinetics and immunogenicity of glycoprotein pharmaceuticals. Detailed characterization of glycans present on recombinant glycoprotein remains an important challenge in the development and production of biotherapeutics. Analytical strategies for characterization of N- and O- glycosylation and monosaccharides analysis will be presented. These include comparison of HILIC-FLR, MALDI-TOF MS and CE-LIF for N-glycan analysis, choice of a method for quantitative and non-selective release of O-linked glycans, and selection of a method for monosaccharide composition analysis. Choosing appropriate glycoanalysis methods allowed detecting changes in glycosylation parameters. A case study will be presented that highlights glycoanalysis techniques useful for gaining understanding of the relationship between process inputs (raw materials) and product quality attributes. The findings confirm that the glycosylation profile of therapeutic antibodies needs to be monitored through development in order to ensure consistency, efficacy, and safety of therapeutic products.

Biography:

Maria Longas completed her PhD at New York University in 1978, and did her postdoctoral training at Columbia University School of Medicine with Dr. Karl Mayer. She has an MA in Organic Chemistry from New York University (1973) and a BA in Chemistry from Hunter College (1971). She is a full Professor of Chemistry at Purdue University Calumet in Hammond, IN. She has more than 24 papers published in reputable journals, and served as a reviewer for several journals.

Abstract:

Ultraviolet light B (UVB) is a technique used to investigate organic molecules. It was utilized in this study to analyze the content of vitamin D3 in face lifted female skin of subjects aged 43, 47, 51, 58, 60 and 66 years. The skin of these subjects had pigmentation of great similarity. It was kept at -40O C, defrosted to room temperature for 2 min and placed in an oven for another 2 min at 105o C. After these 2 min, we took the first UVB reading; a 2nd UVB run was done at room temperature. The amount of vitamin D3 was determined under standard conditions of 7-dehydrocholesterol (7-Dchol). The best temperature to measure D3 was when skin was at room temperature for 15 min. The D3 analysis of the UVB plots of human skin vs. the standard showed a linear decrease in human skin as compared to 7-Dchol. Because 7-Dchol is a direct precursor of vitamin D3, it appears from these results that as the skin ages, 7-Dchol concentration decreases which results in less production of vitamin D3. Conditions to increase vitamin D3 in the aged should be considered, to avoid problems of arthritis, tuberculosis and autoimmune disorders, etc.

Biography:

For the past 35 years, Dr. Szewczuk is Full Professor of Immunology and Medicine, Queen’s University, Kingston, Ontario Canada. He received his B.Sc. in Chemistry (U. of Guelph), M.Sc. in Biochemistry (Guelph), Ph.D. in Immunochemistry (U. of Windsor) and post-doctoral training with Gregory W. Siskind, M.D. in cellular immunology at Cornell University Medical College, NYC. Dr. Szewczuk’s recent research has focused on the role of glycosylation in receptor activation with a particular focus of TOLL-like, nerve growth factor Trk, EGFR and insulin receptors. He has discovered a novel receptor-signaling platform and its targeted translation in multistage of tumorigenesis.

Abstract:

Multicellular tumor spheroids are now at the forefront of cancer research, designed to mimic tumor-like developmental patterns in vitro. Tumor growth in vivo is known to be highly influenced by aberrant cell surface specific sialoglycan structures on glycoproteins. Aberrant sialoglycan patterns that facilitate spheroid formation are not well defined. Here, matrix-free spheroids from human breast MCF-7 and pancreatic PANC1 cancer cell lines and their respective tamoxifen (TMX) and gemcitabine (Gem) resistant variants were generated using the RGD induced self-assembly platform. Pretreatment with specific lectins, exogenous neuraminidase dose dependently reduced spheroid volume formation. Oseltamivir phosphate (OP) promoted cell aggregation and compaction into spheroid formation. PANC1 and triple negative breast MDA-MB231 xenograft tumors from untreated and OP-treated RAGxCγ double mutant mice expressed significant higher levels of α-2,3-sialoglycan (SA) than α-2,6-SA. MCF-7 spheroids expressed higher α-2,3-SA to α-2,6-SA ratio. The relative levels of specific sialoglycan structures on the cell surface facilitate avascular 3D multicellular tumor spheroids in vitro as well as in vivo tumors.

Biography:

Michael Strano is currently the Carbon P. Dubbs Professor of Chemical Engineering at MIT. He received his B.S from Polytechnic University in Brooklyn, NY and Ph.D. from the University of Delaware, both in Chemical Engineering. He was a post doctoral research fellow at Rice University in the departments of Chemistry and Physics under the guidance of Nobel Laureate Richard E. Smalley. From 2003 to 2007, Michael was an Assistant Professor in the Department of Chemical and Biomolecular Engineering at the University of Illinois at Urbana-Champaign before moving to MIT. His research focuses on biomolecule/nanoparticle interactions and the surface chemistry of low dimensional systems, nano-electronics, nanoparticle separations, and applications of vibrational spectroscopy to nanotechnology.

Abstract:

Label-free lectin microarrays are a promising approach to rapidly characterize glycoprotein mixtures. However, to date, demonstrations of highly multiplexed label-free lectin microarrays have been limited. Our group uses near-infrared fluorescent single-walled carbon nanotubes to design glycan-responsive sensors capable of massive multiplexing and real-time detection for incorporation into a label-free lectin microarray. We employ two strategies for the design of our carbon nanotube sensors. The first design platform uses a His-tagged lectin that has been tethered to the nanotube via a Cu2+/NTA linker. We have demonstrated responsivity of these sensors to a variety of natural glycoproteins and to neoglycoproteins constructed from streptavidin and biotinylated sugars. Our second detection platform is based on Corona Phase Molecular Recognition (CoPhMoRe), a technology developed by our group at MIT whereby synthetic, non-biological recognition sites are created from the three-dimensional structure of a carbon nanotube and adsorbed heteropolymer. We have developed CoPhMoRe-based sensors for a variety of molecule types including carbohydrates, resulting in the creation of synthetic lectins capable of being incorporated into the label-free microarray. These sensors, along with a binding kinetic model that we developed, are capable of quantitatively characterizing glycoprotein mixtures at a much shorter time scale than existing characterization techniques. This technology has the potential to address longstanding problems in the fields of biopharmaceutical process analytics and medical diagnostics.

Biography:

Prof. Joon Kim has completed his BS and MS in Microbiology from Seoul National University, PhD in Biochemistry from the University of California at Berkeley and postdoctoral study from Harvard Medical School. He is a Professor in the Division of Life Sciences, and the Director of Radiation Safety and Management Center, Korea University, Seoul, Korea. He has published more than 150 papers in reputed journals

Abstract:

Ribosomal protein S3(rpS3) is a component of the 40S ribosomal small subunit but has multiple other extra-ribosomal functions like apoptosis, cell cycle control, DNA repair etc. It has a DNA repair endonuclease activity which is related with various cancers. Recently, we have discovered that this protein is secreted only from various cancer cell lines as a homodimer but not in normal cells. We also confirmed that rpS3 is secreted more into media from the more invasive cancer cell lines. Presently we confirmed that the secreted protein is glycosylated at the Asn 165 residue and point mutation on this site is defective for the secretion. The secretion pathway turned out to be a ER-Golgi dependent pathway. We propose that glysosylated rpS3 could be used as a useful cancer marker.

Biography:

JDM has completed his PhD at Carlsberg Laboratory, before he moved to Danisco, now Danisco/Dupont, where he worked 20 years as scientist and Research Manager and finally Director of Research in Danisco Biotechnology. He moved to the Technical University in 2008 with a Danisco grant to establish a group working on enzymes, hydrocolloids and prebiotic oligosaccharides. He has published 138 publications and 25 Patents.

Abstract:

Recently, significant progress has been made within enzymatic synthesis of biomimetic, functional glycans, including, for example, human milk oligosaccharides. These compounds are mainly composed of N-acetylglucosamine, fucose, sialic acid, galactose, and glucose, and their controlled enzymatic synthesis is a novel field of research in advanced food ingredient chemistry, involving the use of rare enzymes, which have until now mainly been studied for their biochemical significance, not for targeted biosynthesis applications. For the enzymatic synthesis of biofunctional glycans reaction parameter optimization to promote “reverse” catalysis with glycosidases is currently preferred over the use of glycosyl transferases. Numerous methods exist for minimizing the undesirable glycosidase-catalyzed hydrolysis and for improving the trans-glycosylation yields. This presentation will provide an overview of the approaches and data available concerning optimization of enzymatic trans-glycosylation for novel synthesis of complex bioactive carbohydrates using sialidases, α-L-fucosidases, β-N-acetylhexosaminidases, and β-galactosidases as examples. The use of an adequately high acceptor/donor ratio, reaction time control, continuous product removal, enzyme recycling, and/or the use of co-solvents may significantly improve trans-glycosylation and biocatalytic productivity of the enzymatic reactions. Protein engineering is also a promising technique for obtaining high trans-glycosylation yields, and proof-of-concept for reversing sialidase activity to trans-sialidase action has been established. However, the protein engineering route currently requires significant research efforts in each case because the structure−function relationship of the enzymes is presently poorly understood.

Biography:

Dr. Menter received his PhD degree in Chemistry from the George Washington University in 1969. He completed a postdoctoral fellowship with Prof. Dr. Theodor Foerster at the Institut fuer physikalische Chemie der Universtiaet Stuttgart, Germany. Subsequently, he was at the University of Alabama, Birmingham, and the VA Medical Center (Atlanta) He currently serves as Research Professor of Biochemistry at Morehouse School of Medicine. Dr. Menter is recognized internationally for his work in the areas of collagen photochemistry and melanin photobiology as pertaining to redox reactivity

Abstract:

Mammalian skin epidermis and dermis are usually considered as separate entities, owing to their different functions. However, there is much interaction between them, and it is even suspected that altered dermis can be a factor in epidermal carcinogenesis. In addition to epidermal carcinogenesis caused primarily by UVB (290 – 320 nm) radiation, UVA wavelengths (320 – 400 nm) can cause changes in dermal architecture and composition that causes a response analogous to wound healing. One hypothesis is that the latter changes may render the overall milieu more favorable for epidermal skin cancer formation. In cases of epidermal pigmentary incontinence, melanin pigment may play an additional role in dermal damages. In the present study, we use collagen’s intrinsic fluorescence properties to investigate the action of UV on two acid-soluble type I collagens in an in vitro system. These involve thermal (i.e. temperature dependence in ground state properties) and photochemical (involving excited state transformations). In further experiments, we are beginning a systematic study of the effect of surrounding extracellular matrix (ECM) on our present results. In a preliminary study (to be published) we have found, somewhat surprisingly, that addition of sodium hyaluronate (HA) has little or no effect on the thus far observed properties. We envision expanding the investigation to include the addition of proteoglycans or at least their component chondroitin sulfates.

Biography:

Yves Durocher is a Research Officer at the National Research Council of Canada since 1995. He obtained his PhD in Biochemistry at the Université de Montréal in 1993. Yves manages the NRC’s Mammalian Cell Culture Section which is composed of 33 scientists involved in protein expression and CHO cell line development for internal projects and external clients. His research activities have been focused on the development the of large-scale transient gene expression (LSTGE) platforms using HEK293 and CHO cells for protein production and on the development of stable CHO pool and clonal cell line platforms for the manufacturing of recombinant therapeutic proteins. He also contributed to ~100 scientific publications in peer-reviewed journals.

Abstract:

IgGs that possess Fc-glycans with terminal sialic acid (SA) are thought to be responsible for the anti-inflammatory properties of intravenous immunoglobulins (IVIGs) through a mechanism that is still unclear. The impact of this sialylation on IgG’s effector functions (ADCC and CDC) also remains to be elucidated. To better understand the biological impact of IgG sialylation, there is a need to produce recombinant IgGs with well characterized and more homogeneous glycan structures. The type of SA (NANA or NGNA), the nature of its linkage with the galactose residue (alpha-2,3 or alpha-2,6) or the number of glycan antennae being sialylated, may vary according to the IgG subtype, the host cell in which it is expressed and the cell culture environment. In this study, we show that the a 2,6-sialylation of IgG1’s Fc domain can be efficiently achieved by the transient co-expression of the human beta 1,4-galactosyltransferase-1 (GT) and 2,6-sialyltransferase-1 (ST6) in CHO cells. The process allows for the production of milligram amounts of human-like sialylated monoclonal antibody within two weeks. The impact of this sialylation on IgG1 binding to FcγRIIIa is also presented.

Biography:

Dr. Kubick is head of the department “Cell-free Bioproduction” at the Fraunhofer IZI-BB. His laboratory exploits cell-free protein synthesis as a versatile tool for membrane protein and glycoprotein synthesis, as well as chip-based protein synthesis and translational regulation. In collaboration with Qiagen GmbH he developed novel eukaryotic in vitro translation systems. Dr. Kubick is also a lecturer at the Free University of Berlin and the University of Potsdam. He is an affiliate of the Technical University of Berlin and Lecturer at the University of Applied Sciences, Berlin, Germany.

Abstract:

Membrane proteins and glycoproteins have become an important focus of the current efforts in structural and functional genomics. In contrast, the molecular analysis of posttranslationally modified proteins lags far behind that of non-modified, cytosolic and soluble proteins. Preparing high quality samples of functionally folded proteins represents a major bottleneck that restricts further structural and functional studies. Cell-free protein synthesis systems, in particular those of eukaryotic origin, have recently been developed as promising tools for the rapid and efficient production of a wide variety of posttranslationally modified proteins. This is mainly due to the properties of cultured eukaryotic cells, which are able to carry out many types of posttranslational modifications such as the addition of N- and O- linked oligosaccharides, but also palmitoylation, myristylation, and phosphorylation. Based on these versatile properties of cultured cell lines, we have developed a technique for the standardized production of translationally active eukaryotic cell lysates. Our homogenization procedure avoids any serious breakdown of membrane vesicles already existing in the cytoplasm of the prepared eukaryotic cells. We have demonstrated the functional integrity of these subcellular components by showing signal peptide cleavage as well as glycosylation of in vitro produced proteins. Moreover, we have expanded our cell-free protein synthesis system by the insertion of orthogonal tRNA/synthetase pairs to facilitate the cotranslational and site directed incorporation of non-canonical building blocks. These fluorescently labeled and chemoselective moieties enable the site-specific modification of de novo synthesized glycoproteins.

Biography:

Cheng Ma has completed his PhD from Beijing Proteome Research Center (BPRC). His work as a research scientist in Georgia State University focused on development of novel techniques to analyze the sequence of glycopeptides through the fragment ions of MS/MS. He has a broad background in proteomics, glycoprotemics, glycomics, as well as bioinformatics. Meanwhile, He has more than eight years’ experience in mass spectrometry operation.

Abstract:

Core-fucosylation (CF) plays important roles in regulating biological processes in eukaryotes. Alterations of CF-glycosites or CF-glycans in bodily fluids correlate with cancer development. Therefore, global research of protein core-fucosylation with an emphasis on proteomics can explain pathogenic and metastasis mechanisms and aid in the discovery of new potential biomarkers for early clinical diagnosis. In this study, a precise and high throughput method was established to identify CF-glycosites from human plasma. We found that alternating HCD and ETD fragmentation (AHEF) can provide a complementary method to discover CF-glycosites. A total of 407 CF-glycosites among 267 CF-glycoproteins were identified in a mixed sample made from six normal human plasma samples. Among the 407 CF-glycosites, 10 are without the N-X-S/T/C consensus motif, representing 2.5% of the total number identified. All identified CF-glycopeptide results from HCD and ETD fragmentation were filtered with neutral loss peaks and characteristic ions of GlcNAc from HCD spectra, which assured the credibility of the results. This study provides an effective method for CF-glycosites identification and a valuable biomarker reference for clinical research.

Biography:

Wanyi Guan has completed her PhD from Shandong University, China. She is currently a postdoc in Dr. Peng George Wang’s group in Department of Chemistry in Georgia State University. Her research focuses on enzymatic and chemo-enzymatic synthesis of sugar nucleotide, glycans and their analogs.

Abstract:

In nature, glycans are displayed universally at the surface of living cells. They play important roles in numerous biological events through binding with corresponding glycan binding proteins (GBPs). Usually, the carbohydrate moiety involved in these bindings is composed of the sugar residues at the non-reducing terminal by linked to each other in certain manners and forming specific epitope determinants. Nevertheless, internal epitopes may also participate in the binding and initiate subsequent signaling process. To investigate whether the internal epitope involved in the glycan binding process, an array of symmetric bi-antennary N-glycans were synthesized with tandem epitopes, including LacNAc (Galβ1,4-GlcNAc), sialyl LacNAc (Siaα2,3-LacNAc), 6-sialyl LacNAc (Siaα2,6-LacNAc), Lewis x (Galβ1,4-(Fucα1,3-)GlcNAc), sialyl Lewis x (Siaα2,3-Galβ1,4-(Fucα1,3-)GlcNAc), α-Gal (Galα1,3-LacNAc), and disialic acid (Siaα2,8-Sia), by enzymatic extension of N-glycan in the glycopeptide isolated from chicken egg yolk. For rapid production of glycans, one-pot multiple enzyme (OPME) strategy was employed, and the produced glycans were separated by high performance liquid chromatography monitored with UV detector. In total, 36 glycans were prepared to milligram scale and over 98% purity. Their binding profile to selected GBPs and viruses showed that internal glycan epitopes and modification of terminal epitopes exhibited obvious, but diverse effects to the binding of terminal epitopes.

Biography:

Clifford Lingwood completed his PhD at the University of London at 25 years of age, and postdoctoral studies at the Universities of Washington and Toronto. He has been a professor at the University of Toronto since 1997 and is a senior scientist within the Molecular Structure and Function program of the Research Institute at the Hospital for Sick Children, Toronto He has published more than 200 papers in reputed journals.

Abstract:

Glycosphingolipids (GSLs) accumulate together with cholesterol in membrane lipid rafts, dynamic domains of increased order, which provide foci for transmembrane signaling, membrane trafficking and portals for microbial pathogens. Membrane cholesterol can also complex with GSLs to alter the carbohydrate from a membrane perpendicular to parallel conformation and thereby restrict ligand access to GSLs. Cholesterol ligand access is also restricted in the GSL complex. Since statins are widely used to decrease serum cholesterol levels, we question whether cholesterol depletion affected GSL synthesis. While we found that statins are unable to reduce cholesterol in serum cultured cells, statins nevertheless had a remarkable effect on cell GSL synthesis in that glucosyl ceramide was markedly increased and downstream GSLs increased according to cell line. The enzyme glucosyl ceramide synthase was mislocalized within statin treated cells. This correlated with a loss of cholesterol accumulation in the transGolgi network. The trihexoside, lactotriaosyl ceramide, was induced in 70% of cell lines. This effect was duplicated by the Rab prenylation inhibitor, 3-PEPHC and reversed by addition of the isoprenoid precursor, geranylgeranyl pyrophosphate, a downstream product of HMG Co-A reductase, which is inhibited by statins. These results are consistent with statin induced depletion of Rab prenylation, subsequent aberrant Golgi vesicular traffic, mislocalization of GSL anabolic enzymes and remodeling of GSL synthesis. This process also inhibits the normal retrograde transport of GSLs and the internalization of cholera and Shiga toxins, which hijack GSL retrograde transport for endoplasmic reticulum targeting, are also mislocalized intracellular, thereby protecting statin treated cells.

Biography:

Amélia Pilar Rauter has made her Habilitation at Faculty of Sciences, University of Lisbon (FCUL, 2002) and completed her Ph.D. at Technische Universitaet Graz, Austria (1982). She is the Head of the Carbohydrate Chemistry Group - Center of Chemistry and Biochemistry (FCUL), and has been serving as Editor (RSC Carbohydrate Chemistry book series), Associated Editor (Mediterranean Journal of Chemistry), and member of journals advisory board in Organic/Carbohydrate Chemistry. She is Secretary of the European Carbohydrate Organisation and Secretary of the IUPAC Division on Organic and Biomolecular Chemistry, has published more than 130 papers and book chapters, and authored 12 patents.

Abstract:

The search for new antimicrobial drugs is currently one of the major ongoing research areas due to the spread of multidrug-resistance, thus encouraging research on antibacterial agents with new mechanisms of action. A new family of antibiotics, structurally based on glycolipids, with a potent antimicrobial activity against Bacillus species will be disclosed. We present our recent findings that cover synthetic approaches to new antimicrobial molecular entities differing in the glycon structure, based on alkyl deoxy pento- and hexopyranosides belonging to the D- or L-series. Stereochemistry, in particular the anomeric configuration, and the deoxygenation pattern are modified in order to evaluate their contribution to the structure/activity relationship. Also the lipophilic chain is changed for the recognition of the key structural features for the bioactivity/selectivity for Bacillus species. The surface properties of the most active compounds, in terms of adsorption and aggregation parameters, are also presented. Although surface activity is required for the bioactivity, this property is not sufficient to account for bioactivity, as shown by some promising surface active glycosides, that were not active. In order to have insights into the mechanism of action of this family of compounds, expected to target bacterial membranes, a multidisciplinary approach covering biophysical and biological methods was followed. The elucidation of the new mechanisms of action of this family of antibiotics will be disclosed and discussed, demonstrating the relevance of the glycan structure for the bactericidal activity.

Biography:

Francesco Peri (age: 47) is Professor of Organic and Medicinal Chemistry at the University of Milano Bicocca. He has also a permanent professorship in Medicinal Chemistry at the Ecole Normale Superieure (ENS) in Lyon, France. He coordinates the H2020-funded Marie Curie ETN project TOLLerant: “Toll-Like Receptor 4 activation and function in diseases: an integrated chemical-biology approach” (4 years project, 2015-2019, 8 academic and 2 industrial partners, 13 enrolled PhDs). He has published more than 87 papers in reputed journals, H-index: 24; H-index from 2010: 20.

Abstract:

Toll-like Receptors (TLRs) activation by pathogen associated molecular patterns (PAMPs) is a pivotal molecular event in inflammation and innate immunity, and TLRs and their agonists are responsible for the efficacy of almost every vaccine. Converserly, TLRs hyperactivation by endogenous factors such as oxidized phopsholipids or heat shock proteins is the main cause of many inflammatory and autoimmune diseases. Activating or inhibiting specifically TLRs provides access to a new generation of therapeutics. We developed synthetic molecules able to modulate TLR4 activation and signalling and we studied the mechanism of action (MOA) of these non toxic, drug-like compounds. Positively or negatively charged synthetic glycolipids are active in blocking TLR4 activation by specifical targeting the CD14 co-receptor. These molecules are very efficient in inhibiting TLR4 activation in cells and in contrasting diseases related to TLR4 hyperactivation by infectious and endogenous agents in animal models. We investigated at a molecular level the MOA of these molecules by binding experiments with purified CD14, MD-2 and TLR4 receptors, and experiments on dendritic cells, macrophages and HEK-TLR4 cells. The unique MOA of these molecules is based on the capacity to dissociate CD14 and TLR4 endocytosis, thus creating an inducible CD14 deficiency at the cell surface. These conditions are expected to antagonize TLR4 signaling more effectively than simply competing with LPS for CD14 and TLR4. Very promising results have been obtained at a preclinical level using these drug hits, and recent data on animal models of atherosclerosis, neuroinflammation, and amyotrophic lateral sclerosis will be reported.

Biography:

Maria Longas completed her PhD at New York University in 1978, and did her postdoctoral training at Columbia University School of Medicine with Dr. Karl Mayer. She has an MA in Organic Chemistry from New York University (1973) and a BA in Chemistry from Hunter College (1971). She is a full Professor of Chemistry at Purdue University Calumet in Hammond, IN. She has more than 24 papers published in reputable journals, and served as a reviewer for several journals.

Abstract:

Ultraviolet light B (UVB) is a technique used to investigate organic molecules. It was utilized in this study to analyze the content of vitamin D3 in face lifted female skin of subjects aged 43, 47, 51, 58, 60 and 66 years. The skin of these subjects had pigmentation of great similarity. It was kept at -40O C, defrosted to room temperature for 2 min and placed in an oven for another 2 min at 105o C. After these 2 min, we took the first UVB reading; a 2nd UVB run was done at room temperature. The amount of vitamin D3 was determined under standard conditions of 7-dehydrocholesterol (7-Dchol). The best temperature to measure D3 was when skin was at room temperature for 15 min. The D3 analysis of the UVB plots of human skin vs. the standard showed a linear decrease in human skin as compared to 7-Dchol. Because 7-Dchol is a direct precursor of vitamin D3, it appears from these results that as the skin ages, 7-Dchol concentration decreases which results in less production of vitamin D3. Conditions to increase vitamin D3 in the aged should be considered, to avoid problems of arthritis, tuberculosis and autoimmune disorders, etc.

Biography:

Dr. Razi is the founder of AccuDava Inc., a biomedical company for personalized cancer chemotherapy (http://www.accudava.com), based on the discovery of two Glycomarkers.  Razi received her PhD in Medical Sciences from Uppsala University-Sweden, in 1995. In her post-doctoral program, at the University of California-San Diego, she discovered a novel glycan-mediated lymphocytes activation mechanism in the immune system. She then joined the Consortium for Functional Glycomics (CFG), at The-Scripps-Research-Institute, where her team successfully developed and launched the world’s largest mammalian glycan microarray in 2011. This unique platform that displays 611 defined glycans has since become available globally for studying glycan-mediated interactions. 

Abstract:

Personalized chemotherapy is an unmet need in cancer treatments. A molecular test that can accurately predict the drug efficacy on a patient would be invaluable in selecting effective treatment strategy.

            We have identified two glycan structures, namely Glycomarker-1 and Glycomarker-2, whose expression levels on the cancer cell surface are associated with responses to chemotherapy with platinum drugs (US patent #7585503, and International pending patent). This finding introduces a new concept, linking glycan cell surface expressions with drug reactivity, and proposes a glycan-mediated mechanism for drug uptake.

            Our initial studies profiling the cell surface glycans, using Flow Cytometry with specific glycan-binding lectins, on three isogenic pairs of ovarian carcinoma cell-lines, consisting of chemosensitive and acquired chemoresistant phenotypes, revealed at least a ten-fold decrease in alpha2-6Sialyl-R motif (Glycomarker-1) on resistant phenotypes compared to sensitive cells. Further studies by fluorescent confocal microscopy, colony forming assay, sialidase treatments, and mass-spectrometry confirmed the association of Glycomarker-1 with drug uptake.

            Using Lectin histochemistry (LHC) on clinical samples proved a feasible assay for Glycomarker-1, tested on 64 human ovarian normal and cancerous tissue sections. The LHC on retrospective ovarian cancer specimens, with a known history of drug-response, correctly predicted drug-responses in 22 out of 27 (81.4%) patients.

            During the studies on the mechanism for drug response, another glycan structure, Glycomarker 2, was identified demonstrating a similar expression pattern to Glycomarker 1. Further studies on Glycomarker-2 suggest an association of the two glycomarkers that would put forward a glycan-mediated mechanism for platinum-drug uptake by cancer cells.  

Biography:

Dr. Desai is the Director of Institute for Structural Biology, Drug Discovery and Development at Virginia Commonwealth University. He earned his PhD from Indian Institute of Technology, Bombay and did postdoctoral studies at Iowa, MIT and Illinois. He was awarded the Established Investigator Award by the American Heart Association and the Distinguished Scholarship Award by Virginia Commonwealth University.

Abstract:

Designing glycosaminoglycan (GAG) sequences that target specific proteins is challenging and not yet achieved. This is a key reason that nature’s library of millions of glycosaminoglycans (GAGs) sequences remains largely untapped. We have developed a very generalizable genetic algorithm-based dual-filter screening strategy that addresses this key hurdle. Our computational algorithm utilizes ‘affinity’ and ‘specificity’ filters and a combinatorial library of 46,656 heparan sulfate (HS) hexasaccharides binding to target protein(s) for identifying ‘highly specific’ HS sequences. Synthesis of these sequences followed by biochemical and biophysical studies help test the computational design predictions. This talk will focus on differential targeting of two highly similar proteins and will present design protocol for discovering high specificity GAG sequences. Considering that GAGs bind to several hundred human proteins, majority of which remain un-characterized with regard to GAG interactions, this work lays the pathway for discovering highly selective GAGs as drugs and/or chemical biology tools.

Biography:

Xueli Li completed her PhD from Leipzig University, Germany. She is a research associate in The Michael J Palmieri Metabolic Laboratory, Children’s Hospital of Philadelphia, Philadelphia, PA. She has more than 8 publications in the area of Glycomics recently.

Abstract:

Protein glycosylation is increasingly recognized as a crucial modulator of protein function, offering a third layer of biological information over genomics and proteomics. Modern tools for analyzing released N-glycans from cells, the glycome, have shown abnormal protein glycosylation in numerous human diseases. We developed a quantification of glycome in cells. Upon reaching 100% confluence, the cells were washed twice with PBS and harvested using a cell scraper. The cells were then pelleted and washed with PBS by centrifugation. Fibroblast pellets were lysed in more than 200 μl PBS, and 200 μg protein from the cell lysate was denature and precipitated with 2× volume of 100% propanol. N-linked and O-linked glycans were released from denatured protein, desalted and permethylated before subject to MALDI-TOF analysis. The quantification of O-linked glycans was achieved by spiking glycans from cell with 25μM of C13-labelled T antigen (m/z 543) and C13-labelled sialylated T antigen (m/z 909). N-linked glycans were quantified using 25μM of C13-labelled Man7GlcNAc2 (m/z 1107). Using this method, we identified abnormal fibroblast glycomes in a number of known patients with congenital disorders of glycosylation and demonstrates cellular glycome as an important tool for diagnosis of these diseases.

Biography:

Dr. Menter received his PhD degree in Chemistry from the George Washington University in 1969. He completed a postdoctoral fellowship with Prof. Dr. Theodor Foerster at the Institut fuer physikalische Chemie der Universtiaet Stuttgart, Germany. Subsequently, he was at the University of Alabama, Birmingham, and the VA Medical Center (Atlanta) He currently serves as Research Professor of Biochemistry at Morehouse School of Medicine. Dr. Menter is recognized internationally for his work in the areas of collagen photochemistry and melanin photobiology as pertaining to redox reactivity

Abstract:

Mammalian collagens exhibit weak intrinsic age- and history-dependent UV fluorescence that. UV radiation can cause longer wavelength fluorescent oxidative bands. The fluorescence of both unirradiated and UVC – irradiated acid-soluble calf skin collagen also reflects considerable autoxidation. We have been studying the ground-and excited state behavior of type I acid – soluble collagens from commercial calf skin and from acid-extracted hairless mouse skin as judged by their temperature and wavelength dependence of their fluorescence properties. As the age-related oxidation products destabilize the overall collagen supramolecular structure, we wondered whether the surrounding extracellular matrix affects the ground- and excited state behavior of the collagen polymer in situ. Added sodium hyaluronate (2:1 ratio) shows little or no effect on fluorescence behavior or Arrhenius – type plots. This latter result may indicate that there is little physical interaction between the collagen telopeptide and HA domains in vitro.

Biography:

Tongzhong Ju received his M.D. degree from the Qingdao Medical College
in1986 and his Ph.D. in Biochemistry from Fudan University Shanghai
Medical School (former Shanghai Medical University, China) in 1994. He
completed his post-doctoral training in Biochemistry/Glycobiology in 1999, worked as a Research Associate (1999-2002) and Research Assistant
Professor (2002-2006) at the University of Oklahoma Health Sciences
Center (OUHSC). Since 2006, Dr. Ju has been an Assistant Professor, and now an Associate Professor in the Department of Biochemistry at Emory University School of Medicine. He has published more than 50 papers in many different reputable journals.

Abstract:

Mucin–type O-glycosylation of Proteins is one of the most abundant protein post-translational modifications, and plays important roles in many biological processes, but the repertoire of mucin-type O-glycans synthesized by cells is difficult to determine. Here we developed a novel technology termed Cellular O-Glycome Reporter/Amplification (CORA), a sensitive and versatile method to profile and amplify mucin type O-glycans in any living cells. Cells took up peracetylated benzyl-α-N-acetylgalactosamine, Benzyl-α-GalNAc(Ac)3 into cytosol where it was de-acetylated to form Bn-α-GalNAc, and Benzyl-α-GalNAc was then transported into the Golgi apparatus where it was converted to a large variety of complex O-glycans (Benzyl-O-glycans) by O-glycosyltransferases; and these Benzyl-O-glycans were finally secreted from cells, allowing easy purification for analysis by HPLC and mass spectrometry (MS). CORA resulted in ~100-1000-fold increase in sensitivity over conventional O-glycan analyses, such as β-elimination-MS and identified a more complex repertoire of O-glycans in more than a dozen cell types analyzed. Thus, the high sensitivity and amplification nature of CORA technology makes it possible to analyze entire human O-glycome and others, and offers new opportunities to identify novel glycan biomarkers for human diseases. Furthermore, CORA technology may shed the light on facilitating the development of similar technologies for analyzing/amplifying other types of cellular glycomes.

Biography:

Jan Tkac received his Ph.D. degree in biotechnology at the Slovak University of Technology (Bratislava, Slovakia) in 2000. He received his postdoctoral appointment at the Linkoping University (2001-2003), Lund University (2003-2006) and Oxford University (2006-2008). Currently, he works at the Institute of Chemistry, Slovak Academy of Sciences in Bratislava. He was/is a recipient of an Individual Marie Curie Fellowship (2003) and an ERC Starting grant (2012). His main research activities cover a microarray platform of detection of glycoproteins by lectins with the aid of nanotechnology. He has published 80+ scientific articles, reviews, and 4 book chapters having 1000+ citations.

Abstract:

Integration of nanotechnology into the field of glycomics can address current limitations of fluorescent glycan and lectin microarrays [1,2]. Moreover, label-free transducing schemes especially when working in an ultrasensitive fashion can be applied in a reliable detection of low abundant disease biomarkers [3-5]. For example, electrochemical impedance spectroscopy applied in our studies allows to detect analyte molecules down to a single molecule level (i.e. aM level) if immobilization architecture of ligands (lectins or glycans) is controlled at nanoscale [6-8]. Utilization of such devices in serological glycoprofile of samples from people having some diseases, in analysis of cancer biomarkers, cancer cell lines and viruses will be discussed. Acknowledgement: The financial support from the Slovak research and development agency APVV 0282-11 is acknowledged. The research leading to these results has received funding from the European Research Council under the European Union’s Seventh Framework Program (FP/2007-2013)/ERC Grant Agreement No. 311532 and from the European Union’s Seventh Framework Program for research, technological development and demonstration under grant agreement No. 317420.

Biography:

Dr. Ming Zhang received the Ph.D. degree (Summa Cum Laude) from University of Gottingen, Germany. She has over a decade experience in infectious diseases, in particular targeting vaccine designs for HIV and influenza viruses. She is an Assistant Professor of Epidemiology at University of Georgia, USA, and leads an interdisciplinary research group towards understanding pathogen diversity and mechanisms for vaccine design purposes. She is also keen in educating the next generation of critical thinkers and innovators in infectious diseases.

Abstract:

N-linked glycosylation is a posttranslational modification that has significantly contributed to the rapid evolution of HIV-1. In particular, enrichment of N-linked glycosylation sites can be found within Envelope variable loops, regions that play an essential role in HIV pathogenesis and immunogenicity. In this presentation, I will describe some strategies we learned through though studies of HIV N-linked glycosylation, on both individual and population aspects, that can be implemented in virus vaccine designs, including for influenza prevention purposes.

Biography:

Robert Sabatini has completed his PhD at the age of 30 years from the University of Alabama at Birmingham and postdoctoral studies from the Netherlands Cancer Institute (Amsterdam). He is an Associate Professor at the University of Georgia in the Department of Biochemistry and Molecular Biology. He has published more than 30 papers in reputed journals.

Abstract:

Kinetoplastids are a group of early-diverged eukaryotes that includes the human parasites Trypanosoma brucei, and Leishmania major. The genome of these pathogens contains a modified DNA base (beta-D-glucosyl-hydroxymethyluracil) consisting of O-linked glucose modification of the thymine base such that the glucose moiety sticks out into the major groove of DNA. The modified base, called base J, is synthesized through the hydroxylation of thymidine by a dioxygenase (JBP) forming hydroxymethyluridine, followed by the transfer of a glucose by the glucosyltransferase enzyme JGT. I will present our most recent studies on the synthesis and function of base J, focusing on its role in regulating RNA polymerase II (RNAP II) transcription termination. The genomes of kinetoplastids are organized into polycistronic gene clusters that contain multiple genes that are co-transcribed from a single promoter. We have localized base J at regions flanking the polycistronic clusters at sites involved in initiation and termination and demonstrated that the loss of J results in increased RNAP II recruitment at promoters and transcription of the clusters. We now show that reduction of base J at termination sites within polycistronic gene clusters leads to readthrough transcription and increased expression of downstream genes, including developmentally regulated genes involved in parasite pathogenesis. The current model is that the glucose moiety of base J provides a steric block for transcription elongation stimulating termination. The role of the GTase JGT in regulating the localization of J at specific sites in the genome will also be discussed.

Biography:

Ying Xu is the "Regents and Georgia Research Alliance Eminent Scholar" Chair Professor in Biochemistry and Molecular Biology Department and the Founding Director of the Institute of Bioinformatics, the University of Georgia since 2003. Before that, he was a senior staff scientist and group leader at Oak Ridge National Laboratory. He received his Ph.D. degree in computer science from the University of Colorado at Boulder in 1991. His current research interests are in cancer systems biology and microbial genomics. He has over 300 papers and five books. He is currently the Editor-in-Chief of IEEE Transaction on Computational Biology and Bioinformatics.

Abstract:

We have recently conducted a computational analysis of transcriptomic data of all enzymes involved in the synthesis of glycans of 20 types of cancer versus matching control tissues. Our analyses revealed a number of highly interesting results, including that these glycans predominantly are involved in balancing hydroxyl radicals produced by the diseased cells and some of their oxidized products drive a number of activities typically associated with cancer initiation and development.

Biography:

KLAUS FIEDLER has completed his PhD at the age of 27 years following a Master of Science with a degree in Biology II (Biochemistry) from EMBL Heidelberg with Prof. K. Simons and the Basel University. He carried through his postdoctoral studies at the Rockefeller Research Laboratories, Sloan-Kettering Institute with Prof. J. Rothman. He held a START research grant from the Swiss National Science Foundation thereafter. His first research projects in biological sciences were followed by medical studies at the University of Basel which led to a Master of Medicine. He has published more than 25 papers in reputed journals, monographs and book chapters.

Abstract:

The modeled Golgi dynamics domain of CHOp24, a member of intra-Golgi cargo receptors, scores highly with a collagen-binding domain and carbohydrate-binding modules and is shown to interact with Wnt8 (wingless 8) with a G=-18.3 kcal/mol. Hybrid N-glycans may provide increasing energy of binding up to -7.1 kcal/mol to simulated p24-Golgi dynamics domain-ligand interaction. Apical transport may require N- and O-glycans and thus the interaction may offer insight on epithelial polarity. Groups of regulated molecules in epithelial polarization may include the sorting machinery of epithelial cells, sorted ligands or both, on top of the intricate regulatory mechanisms in substrate and intercellular adhesion. In further research I have analyzed the cargo receptor VIP36 (Vesicular-integral membrane protein of 36 kDa) for carbohydrate interaction. It has been described by me as a lectin in the endoplasmic reticulum-Golgi intermediate compartment, Golgi apparatus and plasma membrane and later was found implicated in parotid gland secretion and apical transport in MDCK cells. The docking reveals top-interacting carbohydrates of the N-glycan and O-glycan class that encompass N-linked glycans of proteins of high mannose and equally complex type which likely function as sorted ligands in epithelial cells. High affinity binding of the ganglioside GM1 carbohydrate headgroup to VIP36 suggests a linkage with protein and glycosphingolipid apical transfer in epithelial cells. Thus, this fundamental approach predicts that interchangeable glycosphingolipid/protein cargo receptor interaction, which may include some p24 family members in glycan binding, may function in apical transport.

Biography:

Dev Arya earned his B.Sc. (chemistry honors program) from St. Stephen's College, Delhi; and PhD (Bioorganic Chemistry) from Northeastern University, Boston. After spending his postdoctoral years in the labs of Prof. T. C. Bruice (UC Santa Barbara), he joined the faculty at Clemson University. Dr. Arya is a recipient of a National Science Foundation CAREER Award (2002) and the ACS Horace S. Isbell Award of the Division of Carbohydrate Chemistry (2007). He has served as the Program Chair (2004-2008), Chair-Elect (2009-2010) and Chair (2010-2011) of the ACS Division of Carbohydrate Chemistry.

Abstract:

Aminoglycosides inhibit bacterial growth by binding to the A-site decoding region of the bacterial 16s ribosomal RNA (rRNA) within the 30S ribosomal subunit. Previous work has shown that there is approximately a five-fold difference in the affinity of neomycin for the human A-site model and the E. coli model. We have developed a screening assay that rapidly identifies compounds that discriminate between the two model rRNA structures. This approach, coupled with a rapid solid phase synthesis method, has identified active aminoglycosides that show large differences in binding affinity for the E. coli A-site and the human A-site than that of neomycin (~30 fold). The methodology for synthesizing, screening for both ribosomal binding/selectivity and bacterial growth inhibition, and rapid analysis of the data provides a systematic method for identification of bacterial ribosome specific antibacterial that can evade bacterial resistance pathways.

Biography:

Nada Al-Hasawi graduated from the Faculty of Science at Kuwait University in 1995 with a degree of Biochemistry and subsequently an M. Sc. In 1999. She worked first as Scientific Assistant, then as Department Technician and then for 5 years as a Teaching Assistant in the Faculty of Pharmacy at Kuwait University before taking up a scholarship to study for a PhD at the Institute of Cancer Therapeutic at Bradford University, England. Following its completiion in 2008 she was appointed to her present position as Assistant Professor in Departmenmt of Pharmaceutical Chemistry, Faculty of Pharmacy at Kuwait University. Dr Al Hasawi’s current interests focus on the role of polysialic acid in metastatic cancers.

Abstract:

It has been known for several decades that alterations in glycosylation patterns play an important role in the metastasis of cancer cells but thus far few drugs have been developed specifically targeting these molecules. Polysialic acid (PSA) is a developmentally regulated cell-surface glycan consisting of sialic acid monomers attached by α-2,8-glycosidic linkages which, in mammals, is mainly expressed on neural cell adhesion molecule (NCAM). Polysialylated NCAM is abundant in embryonic tissues, and limited to areas of persistence of neuronal plasticity in adults. Up-regulation of PSA has also been reported in highly metastatic cancers where it appears to be associated with tumour progression. In this study we tested the ability of crude extracts of the Ayurvedic medicinal plants, namely Withania somnifera (ashwaghanda) and Bacopa monnieri (Brahmi), to inhibit PSA expression in several human tumour cell lines including TE671 rhabdomyosarcoma, Kelly neuroblastoma, 1321N1 brain astrocytoma and HCT colorectal carcinoma. Highest PSA signals, determined by ELISA, were observed in Kelly cells. Hexane extracts of roots of Withania somniferahttp://glycobiology.conferenceseries.com/, at doses that did not cause any cytotoxicity as determined by MTT assay, were found to cause 20-30% inhibition of PSA expression. To exclude the effect of counteracting constituents, we intend to fractionate the herbal extract to purify and characterise the compounds responsible for this inhibitory activity. We propose a therapeutic approach in which expression of glycans associated with tumour progression and metastasis could be inhibited by prolonged treatment with relatively non-toxic agents that do not target cell survival processes.

Biography:

Prof. Joon Kim has completed his BS and MS in Microbiology from Seoul National University, PhD in Biochemistry from the University of California at Berkeley and postdoctoral study from Harvard Medical School. He is a Professor in the Division of Life Sciences, and the Director of Radiation Safety and Management Center, Korea University, Seoul, Korea. He has published more than 150 papers in reputed journals

Abstract:

Ribosomal protein S3(rpS3) is a component of the 40S ribosomal small subunit but has multiple other extra-ribosomal functions like apoptosis, cell cycle control, DNA repair etc. It has a DNA repair endonuclease activity which is related with various cancers. Recently, we have discovered that this protein is secreted only from various cancer cell lines as a homodimer but not in normal cells. We also confirmed that rpS3 is secreted more into media from the more invasive cancer cell lines. Presently we confirmed that the secreted protein is glycosylated at the Asn 165 residue and point mutation on this site is defective for the secretion. The secretion pathway turned out to be a ER-Golgi dependent pathway. We propose that glysosylated rpS3 could be used as a useful cancer marker.

Biography:

Dr. Modenutti received his Bachelors of Science degree in Genetics from the National University of Misiones, Argentina in 2009. He then moved to Buenos Aires to join the working group of Dra. Silvia Hajos at Immunology Department in Faculty of Pharmacy and Biochemistry of University of Buenos Aires (UBA) for his PhD. He worked in close collaboration with Dr. Marcelo Marti at the Chemical Biology of Faculty of Exact and Natural Sciences of UBA . His Ph.D. thesis focused on Hyaluronic Acid glycomimetic drugs development. He continued his work in the Marti Lab as a postdoc. His current work focuses on the development of bioinformatics tools to prediction of protein carbohydrate complex structures.

Abstract:

Proteins that bind carbohydrates are responsible for numerous important biological functions, such as signal transduction, cell adhesion, among many others. Despite the number of reported structures of protein-carbohydrates complexes (PCCs) is constantly increasing, achieving accurate predictions of the protein-carbohydrate interaction by means of Structural Homology Models (SHM) and ligand-docking remains one of the biggest challenges in Glycobiology. This is due mainly because the residues that form the Carbohydrate Binding Site (CBS) can differ from its ideal binding rotamer in the SHM, which can thereafter significantly affect Docking algorithms performance. In addition, while generally available docking programs work reasonably well for most drug-like compounds, carbohydrates and carbohydrate-like molecules are often problematic, because Force-Fields and Scoring Functions are typically designed to reproduce structures of protein-drug complexes. In this work, we present an integrated approach that combines conformational-space sampling of SHM using Molecular Dynamics simulations (MD) and Docking experiments. In order to obtain the most plausible binding structure of receptor and ligand, clustering analysis to identify different conformations was applied. Finally, Water-Site Bias Docking Method (WSBDM, an Autodock based protocol) was performed to generate a diversity of structures and energy-population parameters are used to rank each one of them. Here, we used human Pulmonary Surfactant-associated protein D (hSP-D) as a case study. The results show that this emerges as a promising tool to build reliable 3D-models, which can then be used for rational design or optimization of glycomimetic drugs.

Biography:

Ting Ting Zhao has completed her PhD at the age of 27 years from University of Chinese Academy of Sciences. She has published 19 papers in the international journals.

Abstract:

Diabetic nephropathy (DN) is one of the major microvascular complications of diabetes. Energy metabolism is altered under diabetic conditions. Decorin is a multifunctional small leucine-rich proteoglycan involved in the autophagy and energetic homeostasis. We investigated the role of decorin and its glycosaminoglycan chain in DN. Both decorin and phosphoralated AMPK were highly expressed in the kidney of an accelerated type 1 DN induced by streptozotocin in uninephrectomized Wistar rats at week 20. In high glucose cultured HK2 cell, glycosaminoglycan-free decorin, generated by mutating Ser4 of the mature protein core into Ala (DCN-S4A), showed more significant role in promoting of AMPK phosphoralation and LC3 II compared with the decorin proteoglycan. These data provides clues that the glycosaminoglycan chain of decorin has a reducing effect on activation of AMPK signaling pathway in DN.

Biography:

Graduated from Food and Biotechnology Department, KyungSung University, Busan, Korea in 2010. Obtained Master of Science from the Molecular and Cellular Glycobiology Lab, Department of Biological Science, Sungkyunkwan University, Suwon, Korea in 2012 and presently Ph.D candidate at the same laboratory under supervisor Prof Cheorl-Ho Kim, Sungkyunkwan University. My research interest is Glycan structure and biosynthesis of glycoproteins, glycoconjugates and glycolipids and NeuGc synthesis by CMP-N-acetylneuraminic acid hydroxylase.

Abstract:

Sialic acid is a generic term used for neuraminic acid and its many derivatives. N-glycolylneuraminic acid (NeuGc) is one of the most common sialic acid types found in most mammals except normal humans. NeuGc is closely linked with various diseases including cancer, infections and immune rejection response during pig-to-human xenotransplantation. Biosynthesis of NeuGc is mediated by a specific hydroxylase, cytidine-5’-monophospho-N-acetylneuraminic acid hydroxylase (CMAH), that converts CMP-NeuAc to CMP-NeuGc. In the present study, we isolated pCMAH house-keeping promoter regions (Ph), which are responsible for transcriptional regulation and which are located upstream of the alternative transcript pcmah-2. Luciferase reporter assays using serial construction of each deleted promoter demonstrated that the Pc promoter was highly active in pig-derived kidney PK15. Ph promoter of pcmah lacked a TATA box, but contained three putative Sp1 binding sites. Mutations of these Sp1 binding sites always resulted in the reduction of luciferase activities in Ph-334. In addition, treatment with mithramycin A (25–100 nM) decreased the luciferase activities of the Ph promoters in a dose-dependent manner. Electrophoretic mobility shift assay analysis revealed that the probes containing each Sp1 binding site bound to Sp1. Taken together, the results indicate that Sp1 bind to their putative binding sites on the Ph promoter regions of pcmah and positively regulate the promoter activity in pig kidney cells.

Biography:

Graduated from Biotechnology Department, DongA University, Busan, Korea in 2014. Presently master-doctor integrated coursed student at the Molecular and Cellular Glycobiology Lab, Department of Biological Science, Sungkyunkwan University, Suwon under supervisor Prof Cheorl-Ho Kim. My research interest is Glycan structure and biosynthesis of glycoproteins, glycoconjugates and glycolipids.

Abstract:

NeuGc is acted as an immune rejection antigen in pig to human xenotransplantation, as it is called as a non-Gal xenoantigen, a next xenoantigen to overcome after elimination of major Gal xenoantigen by knocking out the α-1,3-galactosyltransferase in the pig to human xenotransplantation. In the previous study, we isolated two promoter regions of P1 and P2, which are responsible for transcriptional regulation and located on upstream regions of the two alternative transcripts of 5’pcmah-1 and pcmah-2, respectively. Among them, promoter P2 was demonstrated to be responsible for basal house-keeping expression of the gene (BBRC에정). However, since the intestine tissues is important for the selective expression of the gene in responses to the pathogenic infection in pigs, the intestine specific regulatory mechanism of the gene promoter is the best interest of the pig NeuGc biosynthesis. Then, it is in this study reported that the 5’pcmah-1 promoter containing exon 1a and common ORF region (exon 2 to exon 14) is intestine specific in the pig. From the luciferase reporter assays using serial construction of each deleted promoter, it was demonstrated that promoter P1-1600 region relative to upstream region of 5’pcmah-1 is preferentially active in IPI-2I cells than in PK15 cells, corresponding with both mRNA expression patterns. Both promoters lack TATA box, but contain two Sp1 binding sites overlapped in the P1-260. Each mutation of Sp1 binding sites resulted in the reduction of luciferase activities in P1-542, indicating that in the proximal promoter region, Sp1 binding sites are crucial to regulate the intestine specific level of pcmah expression in the IPI-2I cells. In addition, the treatment with mithramycin A (25 nM to 100 nM) decreased the luciferase activity of P1 promoter in a dose-dependent manner. EMSA analysis revealed that the probes containing each Sp1 binding site bind to Sp1 and Sp3. Taken together, the results indicate that Sp1/3, or Sp1 bind to their putative binding sites on the P1 promoter regions of pcmah gene and positively regulate the promoter activity in pig cells.

Biography:

Graduated from Biological Science Department, Myungji University, Yongin, Gyunggido, Korea in 2014. Presently master-doctor coursed student at the Molecular and Cellular Glycobiology Lab, Department of Biological Science, Sungkyunkwan University, Suwon under supervisor Prof Cheorl-Ho Kim. My research interest is Glycan structure and biosynthesis of glycoproteins, glycoconjugates, glycolipids and anti-inflammatory mrchanism

Abstract:

Gangliosides are sialylated glycosphingolipids which are ubiquitously expressed in mammalian plasma membranes and they play crucial roles in cellular interaction, adhesion, differentiation and apoptosis. To synthesize gangliosides in cells, lactosyl ceramide is processed by different pathways composed of various glycosyltransferases or sialyltranferases. One of these sialyltransferases, GD3 synthase (ST8Sia I), is a key enzyme which controls GD3 biosynthesis from its precursor ganglioside GM3. The steady state level of GD3 expression also depends on other enzymes such as GD2 synthase and GT3 synthase. Thus, the relationship between a ganglioside and its related enzymes is linked to its glycosylation pattern in the cell membrane. The GD3 is expressed weakly in normal tissues, and expressed almost exclusively during development or under pathological conditions such as neuronal disorders. In addition, GD3 promotes tumor progression by influencing cellular proliferation, adhesion and metastasis in malignant tumors. However, GD3 is also known to induce apoptosis by transiently accumulating in the cytosol to contribute to mitochondria damage in the early stages of apoptosis. Fas-mediated apoptosis was also induced by GD3 in lymphoid cells. Therefore, the significance of the biological functions of the ganglioside GD3 in cancer cells is still controversial. Although the disialoganglioside GD3 has been considered to be involved in tumor progression or suppression in various tumor cells, the significance of the biological functions of GD3 in breast cancer cells is still controversial. This prompted us to study the possible relationship(s) between GD3 expression and the metastatic potential of a breast cancer MDA-MB231 cells. The human GD3 synthase cDNA was transfected into MDA-MB231 cells, and G-418 bulk selection was used to select cells stably overexpressing the GD3 synthase. In vitro invasion potentials of the GD3 synthase over-expressing cells (pc3-GD3s) were significantly suppressed when compared with control cells. Expression of intercellular adhesion molecule-1 (ICAM-1; CD54) was down-regulated in the pc3-GD3s cells and the decrease in ICAM-I expression is directly related to the decrease in invasiveness of the pc3-GD3s cells. Then, we investigated signaling pathways known to control ICAM-1 expression. No difference was observed in the phosphorylation of ERK and p38 between the pc3-GD3s and control cells (pc3), but the activation of AKT was inhibited in pc3-GD3s, and not in the control (pc3). In addition, the composition of total gangliosides was changed between control (pc3) and pc3-GD3s cells, as confirmed by HPTLC. The pc3-GD3s cells had an accumulation of the GD2 instead of the GD3. RT-PCR results showed that not only GD3 synthase, but also GM2/GD2 synthase (β4-GalNc T) expression was increased in pc3-GD3s cells. Overexpression of GD3 synthase suppresses the invasive potential of human breast cancer MDA-MB-231 cells through down-regulation of ICAM-1, which may be influenced by accumulation of the GD2 ganglioiside.

Biography:

Manuel Maestre Reyna completed his PhD in 2011 at the Philipps University Marburg, Germany, and after a one year stay as lab leader in the same University, moved to the Academia Sinica, Taiwan, to pursue postdoctoral studies in structural proteins of bacterial biofilms. In the past five year, He has published 10 papers in reputed journals and has been involved in a variety of different projects, involving advanced computational and structural biology techniques.

Abstract:

Vibrio cholerae, the causative agent of cholera, is a gram negative bacterium, which may swim freely, or grow in sessile biofilms associated with abiotic surfaces, zoo-plankton, mollusks, or crustaceans in estuarine and brackish waters. Vibrio cholerae biofilms are involved in many aspects of the pathogen's life-cycle, constituting a possible source of antibiotic resistances, and being very important for intestinal colonization. Particularly during exopolysaccharide- dependent biofilm formation, secreted proteins of the rbm gene cluster, including RbmA, RbmC, and Bap1 are key to biofilm ultrastructure, along with the vibrio polysaccharide (VPS) itself. While RbmC and Bap1 have been linked to biofilm-surface interactions , RbmA surrounds cells within the biofilm, which results in improved mechanical strength, and better biofilm accumulation by a not very well understood mechanism, but possibly involving glycan binding. Furthermore, RbmA has been linked to micro-colony formation during V. cholerae pathogenesis, significantly enhancing the infectivity of the pathogen. Here, we elucidated the RbmA crystal structure, as well as demonstrating glycan binding activity. Furthermore, we observed two distinct proteolytic pathways for RbmA. In the first one, protease mediated post-translational modification yielded a different RbmA isoform, RbmA*, involved in VPS independent cell recruitment. The second pathway, which is phosphate and magnesium dependent, possibly leads to RbmA inactivation, and may be involved in biofilm dispersal.

Biography:

Jose Ignacio Ibeas completed a Biology degree in 1991 at Malaga University in Spain and a PhD at the same University in 1996. His PhD work was focused in studding the yeasts involved in sherry wine production. Then he moved to Purdue University in Indiana to study the antifungal properties for the plant PR protein Osmotin. Since 2003 he is Professor at Pablo de Olavide University in Spain were he is Head of the Molecular Biology Department and Principal Investigator for the Fungal Biotechnology group. He has published more than 30 papers in reputed journals and is collaborating as reviewer for many journals and for the Spanish Evaluation Agency.

Abstract:

Cell wall and secreted proteins are the vehicles for the interaction between fungi and their host. Most of these proteins are glycosylated and this posttranslational modification is essential for their localization and function. Although the role for glycoproteins in these interaction have been studied in fungal animal pathogens for many years, almost nothing is known for phytopathogens fungi. Ustilago maydis has raised as an excellent model for the study of plant-pathogen interactions, and its relation with the maize plant is one of the systems in which studies can be tackled from both plant and pathogen perspective. In the last years our group is focused on the study of the role for N- and O-glycosylation during the maize infection for this fungi. We have identified many of the proteins involved in both process, demonstrate that N- and O-glycosylation are both essential for virulence and identified mutants in different stages across the infection process. So we have shown that mutants for Pmt4 (O-mannosyltransferase) are not able to form appressorium, a structure required for plant penetration, mutants for Gls1 (glucosidase I) can not progress into the plant once the plant has been penetrated and Gas2 (glucosidase II b-subunit) mutants are unable to induce plant tumor formation after a defective progression into the plant. By applying in silico analysis we have identified Pmt4 targets which are essential to complete plant infection. Now we are using 2-D gel analysis to identify cytoplasmic, secreted and cell wall glycoproteins involved in the infection process.

Biography:

Dr. Mario Waespy has studied Chemistry and completed his PhD in Glycobiochemistry in November 2015 at the University of Bremen in the laboratory of Prof. Sørge Kelm. Currently, he is establishing his own research project on Trypanosoma trans-sialidases at the Centre for Biomolecular Interactions Bremen (CBIB).

Abstract:

Trypanosomes are protozoan parasites causing trypanosomiasis in Latin America (Chagas' disease) and Sub-Sahara Africa (sleeping sickness in humans and Nagana in livestock). African Animal Trypanosomiasis (AAT or Nagana), caused by Trypanosoma congolense is a devastating disease in domestic and wild African animals causing an annual loss in agriculture productivity of about 4.5 billion USD due to the death of more then 3 million cattle, whereas 40 millions are estimated to be threatened. Trypanosomes express unusual enzymes termed trans-sialidase (TS). TS are surface-bound enzymes, which catalyse the transfer of sialic acids (Sia) from host cell glycoconjugates to terminal galactose residues on acceptor substrates of the parasite’s surface. Since trypanosomes are unable to synthesise Sia de novo they employ TS to cover their surface with Sia to resist and even manipulate host’s innate and acquired immune system and consequently extend their survival in the different hosts. Several studies have shown that TS play important roles in the pathology of trypanosomiasis in mammalian host and are essential for survival of the parasite also in the insect vector, representing TS as a major virulence factor. Our studies on Trypanosoma congolense TS, revealed 14 active variants exhibiting significantly different enzymatic activities, although these can not be sufficiently explained by amino acid variations at the catalytic centre. Besides the catalytic domain (CD), all TS contain a lectin-like domain (LD), whose biological function still remains unknown. Structural and functional characterisation of Trypansoma congolense TS-LD provided new insight into enzymatic activities and correlated biological functions.