In the investigation of the molecular basis for diabetic complications associated with diabetes mellitus, the most studied process is the nonenzymatic glycation (NEG) of human hemoglobin (HbA). The predominant substrate implicated in NEG is glucose. A bound, transient, ring-opened glucose can be generated in the presence of bound inorganic phosphate (Pi). This Pi-induced ring-opened glucose serves as an electrophile and is able to react with a nucleophilic lysine or N-terminal valine to generate structurally-modified proteins that may induce pathology. However, other Pi-catalyzed electrophilic substrates can be generated in a physiological setting (referred to as Pi-catalyzed glucose degradation products, or GDP). Computational modeling with representative GDP substrates (including glyoxal) predicts that exothermic binding to the β-chain of HbA takes place in sites known to glycate (Val1,Lys8, Lys17, Lys82, Lys132, and Lys144). Glucose binds to identical HbA sites with comparable exothermicities. GDP binding exothermicity and site specificity depend upon the specific GDP and the charge state of amino acid residues. The percent of geometrically favorable binding events of GDP greatly exceeds that of glucose. Thus, Pi-catalyzed GDP formation occurs prior to binding, generating reactive electrophiles that can bind competitively with glucose in known HbA glycation sites. The bound GDP structures possess a higher geometric probability for reaction with protein amino acid residues than glucose.  Moreover, incubation of glyoxal with bovine serum albumin (BSA) indicates that protein crosslinking takes place.  Therefore, Pi can play at least two roles in NEG prior to the covalent stages of the process that can lead to the pathology of diabetes mellitus. Prior to binding, Pi-catalyzed GDP formation can take place producing electrophiles that later bind and can lead to structurally-modified proteins.  Pi can also concomitantly bind with and ring open glucose, generating the electrophilic isomer of glucose that proceeds in NEG. <br/>

Towards an understanding of nonenzymatic glycation #4: The role of phosphate-catalyzed glucose degradation products in non-covalent protein binding

251st National Meeting (2016)

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