The Influence of Aging and Glycation on Protein-Thiol Mixed Disulfides in the Eye Lens




Dickerson Jr., Jaime E.


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Jaime E. Dickerson, Jr., The Influence of Aging and Glycation on Protein-Thiol Mixed Disulfides in the Eye Lens. Doctor of Philosophy (Biomedical Sciences), June, 1994, 163 pp., 9 tables, 28 illustrations, bibliography, 116 titles. The human lens is continually growing. As new cells are formed they differentiate into fiber cells which have no organelles, no protein synthesis or turnover. Lens protein aging involves formation of very large aggregations and insoluble complexes. These are held together through disulfide linkages. Reduced gluthathione (GSH) is present in high concentrations. The oxidized form, (GSSG), (5% of the total) can form mixed disulfides with proteins. This can destabilize the protein conformation. Accumulation of mixed disulfides may increase the potential for further modification. The participation of a PSSG (protein/gluthathione mixed disulfide) in the formation of a protein-protein disulfide becomes increasingly likely. The purpose of this work is to document PSSG and protein-cysteine mixed disulfide (PSSC) accumulation in human lenses (through eight decades), and to identify a third mixed disulfide discovered in this research. The free thiol molecules GSH and cysteine were also quantitated for normal and cataractous lenses. Glycation may alter conformation similar to mixed disulfides and potentiate mixed or protein-protein disulfide formation. This model was evaluated two ways. First, purified alpha crystalline was incubated with ascorbate and conformational changes were evaluated with CD spectroscopy. Second, rat lenses were cultured under high sugar conditions to determine if the resulting glycation influenced the level of mixed disulfides. Conversely, the effect of prior mixed disulfide formation on the extent of glycation in another purified crystalline, gamma, was evaluated. The results indicate: GSH declines in the lens with age, cysteine exists in the lens albeit at relatively low levels, PSSG shows a triphasic pattern of accumulation, PSSC accumulated linearly with increasing age, the existence of a third mixed disulfide species, gamma glutamylcysteine mixed disulfide, detected in old or cataractous lenses, has been confirmed, glycation by ascorbic acid alters α- crystalline secondary structure, the influence of glycation is minimal on mixed disulfide formation, mixed disulfide formation affects glycation of gamma crystalline.