Formation of MethylGlyoxal Induced, Partially De-Aggregated Soluble Advanced Glycation Endproducts of Human Fibrinogen Short Title: Glycation of Human Fibrinogen by MethyGlyoxal

Advanced glycation endproducts (AGEs), a heterogeneous group of molecules, are formed from the non-enzymatic reaction of reducing sugars or their metabolites with free amino acids in plasma or amino groups of proteins, lipids or nucleic acids. AGEs can accumulate in plasma and tissues. Once formed, they can contribute to the pathology of a broad range of conditions including diabetes, Alzheimer’s disease, cataract formation and coronary heart disease. The objective of this project was to demonstrate the in-vitro formation of advanced glycation endproducts by the reaction of human fibrinogen with MethylGlyoxal (MG), a reactive sugar metabolite. Human fibrinogen (hFib) at physiological concentrations was glycated with MG at high, observable concentrations. This showed a time dependent, hyperchromic shift in the UV absorbance at 280 nm, which indicated an increase in electron stacking suggesting an increase in adduct formation. Furthermore, the λmax underwent a hypsochromic shift changing the wavelength from ~280 nm to ~265 nm after 21 days of incubation, further suggesting an increase in adduct formation. Results from HPLC and UV analysis supported the formation of a Partially De-Aggregated Soluble Advanced glycation product, (PDASA). This re-solublized aggregate may be the result of a reversible clot formation model which would be useful in subsequent experiments on the thermodynamics of human fibrinogen conformation changes.