EFFECT OF THE UV MODIFICATION OF ALPHA-CRYSTALLIN ON ITS ABILITY TO SUPPRESS NONSPECIFIC AGGREGATION

Recent studies have shown that structural modifications of alpha-cryst allin during lens aging decrease it's effectiveness as a molecular cha perone. Some of these posttranslational modifications have been linked to UV radiation, and this study was undertaken to investigate the eff ect of UV irradiation on the ability of alpha-crystallin to suppress n onspecific aggregation. The effect of 3-hydroxykynurenine (3-HK) was a lso investigated as a model for its glucoside (3-HKG), a main lens chr omophore that has been linked to photochemical changes in the human le ns. Alpha- and gamma-crystallin solutions (1 mg/mL, 1:0.125 wt/wt) wer e photolyzed (transmission above 295 nm) for various time intervals. T hermal denaturation of gamma-crystallin with or without alpha-crystall in was carried out at 70 degrees C and increases in light scattering w ere measured at 360 nm. We found that (1) irradiation of gamma-crystal lin increased its susceptibility to heat-induced scattering, The addit ion of alpha-crystallin protects it against thermal denaturation, alth ough its ability to do so decreases the longer gamma-crystallin is irr adiated and (2) irradiation of alpha-crystallin decreases its ability to suppress nonspecific aggregation and the presence of 3-HK during ir radiation decreases it further, Our results indicate that posttranslat ional modifications of alpha-crystallin due to UV irradiation affect t he sites and mechanisms by which it interacts with gamma-crystallin, T he kinetics of gamma-crystallin unfolding during thermal denaturation were also analyzed, We found that a simple two state model applies for nonirradiated gamma-crystallin, This model does not hold when gamma-c rystallin is irradiated in the presence or absence of alpha-crystallin . In these cases, two step or multistep mechanisms are more likely.