Dehydroalanine and Dehydrobutyrine: Post-Translational Modifications Implicated in Aging Lenses and Age-Related Nuclear Cataracts

Abstract

The molecular mechanisms associated with aging and age-related diseases are still being discovered. Spontaneous degradation of proteins can lead to the formation of post-translational modifications (PTMs). Among the PTMs found among long-lived proteins, have been those that lead to irreversible protein-protein crosslinks, such as those seen in the ocular lens. Two PTMs that have been implicated in crosslink formation in age-related nuclear cataracts (ARNC) are dehydroalanine (DHA) and dehydrobutyrine (DHB). DHA is formed from cysteine/serine and DHB is formed from threonine residues. Even though DHA and DHB-mediated crosslinks have been found in ARNC, a study of DHA and DHB and how it may contribute to aging and age-related diseases has been the focus of few studies. In this dissertation work, the prevalence of DHA and DHB were studied in aging human lenses with and without ARNC. DHA-containing peptides were classified into three groups based on abundance changes with age and cataract: those that (1) increased only in ARNC, (2) increased in aged and cataractous lenses, and (3) decreased in aged lenses and ARNC. In most donor groups, proteins with DHA and DHB were more likely to be found among urea-insoluble proteins rather than among water- or urea-soluble proteins. To further study whether DHA may induce structural changes, the prevalence of DHA from cysteine loss and similarly DHA-mediated glutathione crosslinks were studied in an established ARNC animal model. DHA abundance in control guinea pig lens proteins was compared to guinea pig lens proteins exposed to a hyperbaric oxygen treatment. The results showed that DHA was more likely to be in the water-insoluble fraction while DHA-GSH crosslinks were more likely to be in the water-soluble fraction. An alanine mutant was used to measure the potential structural differences that may be caused by DHA formation in β B1-crystallin, a lens protein that was found to have DHA from previous results. These results showed that DHA may have a small but measurable impact on protein solubility.