Structure and stability of gamma-crystallins. II. Differences in microenvironments and spatial arrangements of cysteine residues.
The gamma-crystallin fractions II, III and IV from calf eye lens were treated with the thiol-specific fluorescent probe 2-(4'-maleimidylanilino)naphthalene-6-sulfonate (MIANS), in order to determine the reactivity of the seven (gamma-II) or six (gamma-III, gamma-IV) cysteine residues. Two classes of reactive cysteines were distinguished by variations in fluorescence intensity with increasing molar excess of the probe, and approximately three cysteines were nonreactive in each gamma-crystallin. From the position of the emission maximum, it is apparent that MIANS-labeled cysteines of gamma-IV are in the least hydrophobic environment. Fluorescence energy transfer was observed from tryptophan to MIANS-labeled cysteines in both gamma-II and gamma-III crystallins, with efficiencies of 86% and 89%, respectively, but not in gamma-IV crystallin. We suggest that the spatial arrangements and microenvironments of cysteine residues of gamma-crystallins are sufficiently different from each other to account for the variations in fluorescence characteristics of the MIANS-labeled proteins and the lack of energy transfer in gamma-IV crystallins.
D000817:Anilino Naphthalenesulfonates; D000818:Animals; D002417:Cattle; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003459:Crystallins / Q000032:analysis*; D003545:Cysteine / Q000032:analysis*; D008433:Mathematics
D000817:Anilino Naphthalenesulfonates; D003459:Crystallins; C021310:2-(4'-maleimidylanilino)naphthalene-6-sulfonic acid; D003545:Cysteine