1. O-glycosylation of Glycogen Metabolism Enzymes By Onyinyechi Chima-Okereke. Bio 466H

2. Background. Oglycosylation is a post-translational modification. The substrate for O- glycosylation, UDP- GlcNAc, is synthesized in the hexosamine biosynthesis pathway. The dynamic cycling of O-GlcNAc allows for cellular regulation

3. Hypothesis: To determine if any glycogen metabolizing enzymes are o-glycosylated. Method: 1. Extract glycogen pellet from rabbit muscle 2. Digest this glycogen pellet with human salivary amylase. 3. Detect proteins modified by O-GlcNAc using: Standard Western Blotting techniques The lectin sWGA-HRP

4. Expected Results. There is a high concentration of glucose and its metabolites in the glycogen pellet. UDP-GlcNAc can be synthesized through the HBP, thus, allowing for O-GlcNAc modification. Glycogen synthase, glycogen phosphorylase, the branching and debranching enzymes are expected to be o-glycosylated.

5. Relevance. O-glycosylation is involved in the mechanism for the development of diabetes. Streptozotcin (STZ), an irreversibly inhibitor of O- GlcNAcase, leads to an irreversibly massive increase in O-glycosylated protein within the beta cell. Alloxan is an irreversible inhibitor of OGT. It disrupts the ability of beta cells to O- glycoylate intracellular proteins. High levels of glucose, hyperglycemia, causes increased reversible beta cell O-glycosylation.

6. Relevance cont…. This means that in type II diabetes, excessive carbohydrate intake and the development of insulin resistance, is likely to cause increased beta cell o- glycosylation. This hyperglycemia has the same effect on the beta cell as STZ but it is reversible, thus, less toxic. The gradual and chronic beta cell toxicity caused by hyperglycemia induced o- glycosylation induces the very hyperglycemia itself →more beta cell toxicity → downward spiral. With more research, it may be possible to prevent the glucotoxicity of type II diabetes

7. Acknowledgements. Dr. Khatra (Research Mentor). Dr. Mason Howard Hughes Medical Institute.