Abstract— The effect of 300 nm irradiation on the sulfhydryl groups of calf lens a‐crystallin has been investigated by using specific, covalently bound fluorescent sulfhydryl probes 4–(N‐iodoacetoxy)ethyl‐N‐methylamino‐7‐n‐itrobenz‐2‐o‐xa‐1,3‐d‐iazole (IANBD), N‐iodoacetyl‐N′‐(5‐s‐ulfo‐l‐naphthyl) ethylene‐diamine (1,5 IAEDANS) and 5‐i‐odoacetamidofluorescein (IAF). The decrease in tryptophan fluorescence with time of irradiation of a‐crystallin, is accompanied by a decrease in the fluorescence of the hydrophobic sulfhydryl label IANBD. In addition, the fluorescence of the surface‐sulfhydryl label IAF increased in the irradiated a‐crystallin. These results indicate that the sulfhydryl groups are in a more exposed (hydrophilic) environment in the irradiated protein than in the control, possibly because of partial unfolding of the protein. This result is confirmed by fluorescence lifetime measurements with IAEDANS. The decay curve of IAEDANS‐α‐crystallin has a major lifetime of 15.7 ns and a minor one of 24.6 ns. Upon irradiation, the lifetime of the major component decreases to 10.2 ns and that of the minor component to 21.7 ns. Denatured IAEDANS‐α‐crystallin has a single lifetime of 10.4 ns. These results show that the photoinduced damage to the tryptophan residues of α‐crystallin alters the environment of the sulfhydryl groups and induces a change in the tertiary structure of the protein. Proximity of the cysteine residues to tryptophan in the tertiary structure of the protein may be an important determinant of their susceptibility to photoinduced change.
|Number of pages||7|
|Journal||Photochemistry and Photobiology|
|State||Published - Feb 1986|