Distinct interactions of αA-crystallin with homologous substrate proteins, δ-crystallin and argininosuccinate lyase, under thermal stress

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• 作者：Ya-Huei Chen^a ; ¹ ; Ming-Ting Lee^a ; ¹ ; Yu-Wen Cheng^a ; Wei-Yuan Chou^a ; Chung-Ming Yu^b ; Hwei-Jen Lee^a ; ^{hjlee@mail.ndmctsgh.edu.tw}
• 关键词：&#x3b1 ; A-Crystallin ; Molecular chaperone ; Goose &#x3b4 ; -crystallin ; Argininosuccinate lyase ; Thermal stability ; Protein aggregate
• 刊名：Biochimie
• 出版年：2011
• 出版时间：February 2011
• 年：2011
• 卷：93
• 期：2
• 页码：314-320
• 全文大小：879 K

文摘

δ-Crystallin is a taxon-specific eye lens protein that was recruited from argininosuccinate lyase (ASL) through gene sharing. ASL is a metabolic enzyme that catalyzes the reversible conversion of argininosuccinate into arginine and fumarate and shares about 70 % sequence identity and similar overall topology with δ-crystallin. ASL has a lower thermal stability than δ-crystallin. In this study, we show that the small heat shock protein, αA-crystallin, functions as a molecular chaperone, and enhanced thermal stability of both δ-crystallin and ASL. The stoichiometry for efficient protection of the two substrate proteins by αA-crystallin was determined by slowly increasing the temperature. N- or C-terminal truncated mutants of δ-crystallin co-incubated with αA-crystallin showed higher thermal stability than wild-type enzyme, and the stoichiometry for efficient protection was the same. Thermal unfolding of δ-crystallin or ASL in the presence of αA-crystallin followed a similar three-state model, as determined by circular dichroism analyses. A stable intermediate which retained about 30 % α-helical structure was observed. Protection from thermal denaturation by αA-crystallin was by interaction with partly unfolded ASL or δ-crystallin to form high molecular weight heteroligomers, as judged by size-exclusive chromatography and SDS-PAGE analyses. Aggregate formation of ASL was significantly reduced in the presence of αA-crystallin. The extent of protection of ASL and δ-crystallin at different ratios of αA-crystallin were described by hyperbolic and sigmoidal curves, respectively. These results suggest the preferential recognition of partly unfolded ASL by αA-crystallin. In contrast, unstable δ-crystallin might trigger a cooperative interaction by higher stoichiometries of αA-crystallin leading to fuller protection. The different interactions of αA-crystallin with the two homologous but functionally different substrate proteins show its behavior as a chaperone is variable.