In diabetes, the accumulation of sorbitol is believed to be a key factor in initiating cataract, retinopathy, and neuropathy. We are interested in controlling the accumulation of sorbitol by regulating the action of the two enzymes of the sorbitol pathway: (1) aldose reductase (AR), which reduces glucose to sorbitol, and (2) sorbitol dehydrogenase (SDH), which oxidizes sorbitol to fructose.
Our aim i s to design innovative methods to inhibit the action of AR or increase SDH, with the purpose of reducing sorbitol accumulation in diabetic tissues. Site-directed mutagenesis of AR has been a major priority of our laboratory. We have made amino acid substitutions in the rat and human AR and determined that some of these changes to affect the kinetics of the protein with its substrate. For example, when histidine glutamine at position 110 was changed to glutamine, the activity of AR was reduced dramatically. The H110Q mutant protein showed very little activity with glyceraldehyde (1% of normal) and no activity with rho-nitrobenzaldehyde. Other histidine substitutions did not acutely alter the kinetics of AR, supporting the finding that H110 plays an essential role in catalysis. These structure/function studies should help define the active site and locate the target areas of the current AR inhibitors. Another strategy to control sorbitol accumulation is to regulate SDH. We have determined the primary sequence of human SDH and characterized part of the SDH gene. Molecular genetic studies also are under way to test for an SDH genetic defect in a family presenting with congenital cataracts and lowered SDH enzyme activity. By evaluating the expression of SDH at the gene level, we may be able to evaluate its role in sorbitol accumulation in diabetes and other genetic diseases.
