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STRUCTURE AND STABILITY OF γ‐CRYSTALLINS‐IV. AGGREGATION AND STRUCTURAL DESTABILIZATION IN PHOTOSENSITIZED REACTIONS
Author(s) -
Mandal Krishnagopal,
Kono Masahiro,
Bose Swapan K.,
Thomson John,
Chakrabarti Bireswar
Publication year - 1988
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1988.tb08848.x
Subject(s) - chemistry , singlet oxygen , photochemistry , iodoacetamide , tryptophan , reaction rate constant , cysteine , methylene blue , fluorescence , protein aggregation , guanidine , crystallin , oxygen , biophysics , kinetics , amino acid , biochemistry , organic chemistry , physics , quantum mechanics , photocatalysis , biology , enzyme , catalysis
— Unlike α‐ and β‐, γ‐crystallins become turbid upon irradiation with 300 nm or white light in the presence of photosensitizers, e.g., methylene blue (MB) or riboflavin (RF). These proteins, however, do not aggregate in the presence of guanidine hydrochloride. Turbidity formation is concentration‐dependent. Except in RF‐sensitized reactions, the onset and rate of turbidity formation are faster in γ‐IV than in the other two crystallins. Labeling the thiol groups of the protein with iodoacetamide does not change the turbidity rate in case of irradiation with 300 nm light, but it does change it in case of RF‐ or MB‐sensitized reaction. The order of rate constants of the decrease in tryptophan emission under aerobic conditions upon 300 nm irradiation and MB‐ and RF‐sensitized reactions is γ‐IV > γ‐III > γ‐II. The rate constants in the absence of air and in the presence of D 2 O upon MB‐ and RF‐sensitized reactions also indicate that the role of singlet oxygen is significant in the former reaction. We suggest that the photoinduced structural changes occur in two steps. In the first, photooxidation of tryptophan as well as cysteine residues (including the buried cysteines) occurs, leading to small conformational changes of the protein. Conformational changes of the protein, as evident from the near‐UV CD and fluorescence lifetime measurements, subsequently result in aggregation of the protein. As suggested by X‐ray analysis, the perturbation of the cys 78 and 32 by the active species of oxygen appears to be responsible for the destabilization of the protein structure, resulting in rapid aggregation of these crystallins.