NEW METABOLITES AND PATHWAYS IN THE MAMMALIAN LENS

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We have identified a novel phosphomonoester fructose 3-phosphate (Fru-3-P) in the diabetic rat lens and shown it to be a potent crosslinking agent of crystallins in vitro. We have also demonstrated that Fru-3-P decomposes to 3-deoxyglucosone (3dG) a potent protein crosslinker, and have shown that this dicarbonyl is present in the diabetic lens. In this research program we propose to further assess the importance of Fru-3-P in the diabetic rat lens by identifying other possible glycating agents in this tissue and to evaluate their glycation potential relative to Fru-3-P. If Fru-3-P turns out to be the dominant glycating agent, as suggested by preliminary data, we will subsequently determine the kinetic parameters of Fru-3-P and 3dG mediated glycation of rat crystallins in vitro and determine the amount glycation mediated by these compounds in the diabetic rat lens in vivo. Since fructose 3-phosphate is synthesized from fructose by a novel enzyme fructose-3-phosphokinase, we will purify this enzyme and determine its molecular identity as well as its kinetic and mechanistic properties.