Exploring the Biological Roles of Glycans Max Nanis

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Principles & Conclusions about Biological Roles of Glycans Range complete importance spectrum Enormous diversity Serve as a protein attachment surface War of microbes is fought on a battlefield of carbohydrates!

Consequences of Altered Glycosylation Variate Highly variable (intra class) - studied via: Prevention of initial glycoslation Alteration of oligosaccharide processing Enzymatic or chemical deglycoslation of completed chains Genetic elimination of glycosylation sites

Structural & Modulatory Roles Protective, stabilizing, organizational, barrier functions, chaperonin Modulate interaction with one another. GF receptors binding mechanism Some specificity required by defined oligosaccharide sequence Tuning: Heparin/heparan sulfate chains on anticoagulant antithrombin III, tyrosine phosphorylation acitivty of EGF receptor & insulin receptor

Specific Ligands for Exogenous Receptors Extrinsic GBPs Specific receptors for viruses, bacteria, parasites and symbiotic relationships Some glycan sequences act as decoys for microoranisms and parasites ex. Mucosal cell decoys with specific oligosaccharide ligand in mucin, then are washed away

Specific Ligands for Endogenous Receptors Intrinsic GBPs Earliest known mediated clearance, turnover and intracellular trafficking Free oligosaccharides may induce highly structure-specific hormonal responses Modern: cell- cell recognition, cell- matrix interactions ex. Sperm:Egg binding via O-glycans

Homogeneous Glycans Can Have Different Roles Expression of specific types of glycosylation of different glyconjugates in different tissues at different times of development implies these structures have diverse roles in similar organism. New oligosaccharide or modification can evolve independently in different tissues and at a different time in development.

Universal Roles? Species-specific variation in glycan structure Different selection pressures via pathogen exposure

Structure Variation What components of glycans mediate essential biological roles? Terminal sequences, modifications, and unusual structures

Determining Biological Roles Structure and biosynthesis doesn’t imply understanding of function Many functions discovered by chance Difficult to differentiate between functions mediated by each glycan

Lectins / Antibodies Lectins (sugar-binding proteins) and antibodies that are glycan specific Localization (S35?) Not reliable when introduced into systems where cross-creating glycan structures are potentially present (In vivo?) Nonspecific aggregation

Lectins / Antibodies

Metabolic Inhibition or Alteration of Glycosylation Pharmacological agents can inhibit glycosylation Good to determine pathway Con: May degrade membranes

Metabolic Inhibition or Alteration of Glycosylation

Natural Glycan Ligands:Specific Receptors Promote potential lectin in high enough quantities, can look for specific ligands Red cell agglutination, flow cytometry, surface plasmon resonance, and affinity chromatography Con: Affinity of lectin for ligand may be low (need higher densities)

Natural Glycan Ligands:Specific Receptors

Receptors Recognizing Specific Glycans Search for specific receptor Hemagglutination, flow cytometry, and affinity chromatography Reasonable quantities of purified glycan Con: Lectin interactions can be low affinity, thus should use multivalent form of glycan Most relevant receptor may be found in another organism!

Receptors Recognizing Specific Glycans

Interference by Soluble Glycans or Structural Mimics SG or SM can interfere with interaction between lectin and glycan Con: inhibitor cross-reacting with with unknown receptors

Interference by Soluble Glycans or Structural Mimics

Glycan Structure Elimination Glycosidases are degradative and highly specific for certain glycan sequences Does not interfere with biosynthetic machinery, but selectively eliminates structure (post synthesis)

Glycan Structure Elimination

Glycan Mutants Con: defects in glycosylation may have limited/not easily discernable consequences Often affect multiple systems/unpredictable phenotype Redundant or fail-safe pathways possible Best to analyze structural functions Genetic defects relatively uncommon

Glycan Mutants

Receptor Mutants Receptor alteration can yield informative phenotype in regards to function Con: receptor protein my have unrelated functions to glycan recognition

Receptor Mutants

Functions Protection: Cushions the plasma membrane and protects it from chemical injury Immunity to infection: Enables the immune system to recognize and selectively attach foreign organisms Defense again cancer: Changes in the glycocalyx of cancerous cells enable the immune system to recognize and destroy them

Functions Transplant compatibility: Forms the basis for compatibility of blood transfusions, tissue grafts, and organ transplants Cell adhesion: Binds cells together so that tissues do not fall apart Inflammation regulation: Glycocalyx coating on endothelial walls in blood vessels prevents leukocytes from rolling/binding in healthy states

Functions Fertilization: Enables sperm to recognize and bind to eggs Embryonic development: Guides embryonic cells to their destinations in the body