THE GENERATION OF SUPEROXIDE ANION BY THE UVA IRRADIATION OF HUMAN LENS PROTEINS

In this study we report the generation of superoxide anion and hydroge n peroxide by a water-insoluble protein fraction from aged human lense s in response to WA light. Irradiation with 1.5 kJ cm(-2) of UVA light (> 338 nm) over a 1 hr period caused the formation of 20 +/- 0.1 mu M superoxide radical and 37 +/- 0.5 mu M hydrogen peroxide. A linear ph otolysis of SH-groups (21 nmol ml(-1), 26%), His (117 nmol ml(-1), 26% ) and Trp (72 nmol ml(-1), 27%) residues was seen following 60 min of irradiation. The addition of SOD, however, had no effect on the photol ytic destruction of any of these amino acid residues. Incubation of th e human WISS proteins and bovine alpha-crystallin in the presence of 4 3-49 mu M of O-2(-) generated in a xanthine oxidase/hypoxanthine syste m over a 1 hr period, caused no loss of histidine, little or no loss o f tryptophan and loss of 7-9 nmol ml(-1) of sulfhydryl groups with bot h proteins. This argues that O-2(-) can only account for the destructi on of at most 4-8 nmol SH-groups in human water-insoluble proteins fol lowing 1 hr of UVA irradiation. (C) 1996 Academic Press Limited