Tryptophan Cluster Protects Human D-Crystallin from Ultraviolet Radiation-Induced Photoaggregation In Vitro

Exposure to ultraviolet radiation (UVR) is a significant risk factor for age-related cataract, a disease of the human lens and the most prevalent cause of blindness in the world. Cataract pathology involves protein misfolding and aggregation of the primary proteins of the lens, the crystallins. Human gamma D-crystallin (H gamma D-Crys) is a major gamma-crystallin in the nucleus of the human lens. We report here analysis of UVR-induced damage to H gamma D-Crys in vitro. Irradiation of solutions of recombinant H gamma D-Crys with UVA/UVB light produced a rise in solution turbidity due to polymerization of the monomeric crystallins into higher molecular weight aggregates. A significant fraction of this polymerized protein was covalently linked. Photoaggregation of H gamma D-Crys required oxygen and its rate was protein concentration and UVR dose dependent. To investigate the potential roles of individual tryptophan residues in photoaggregation, triple W:F mutants of H gamma D-Crys were irradiated. Surprisingly, despite reducing UVR absorbing capacity, multiple W:F H gamma D-Crys mutant proteins photoaggregated more quickly and extensively than wild type. The results reported here are consistent with previous studies that postulated that an energy transfer mechanism between the highly conserved pairs of tryptophan residues in H gamma D-Crys could be protective against UVR-induced photodamage.