Differential temperature-dependent chaperone-like activity of αA- and αB-crystallin homoaggregates
Differential temperature-dependent chaperone-like activity of αA- and αB-crystallin homoaggregates
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DOI：
10.1074/jbc.274.49.34773
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发表时间：
1999-12-03
影响因子：
4.8
通讯作者：
Rao, CM
中科院分区：
文献类型：
--
作者：
Datta, SA;Rao, CM
Alpha-Crystallin是一种由αA-和αB-晶体蛋白组成的异多聚体蛋白，其功能是作为分子伴侣阻止蛋白质聚集。我们早些时候已经证明，α-晶体蛋白的结构扰动可以几倍地增强其伴侣样活性。α-晶体蛋白的两个亚基具有广泛的序列同源性，并各自显示出伴侣样活性。我们研究了αA-和αB-晶体蛋白均聚体对热和非热聚集模式的伴随性。我发现，对于非热聚集模式，αB-晶体蛋白即使在亚生理温度下也显示出显著的保护能力，在这种温度下，αA-晶体蛋白或异多聚体α-晶体蛋白表现出很少的伴侣样活性。有趣的是，这些均聚体对β(L)-晶状体蛋白热聚集的保护能力的差异可以忽略不计。为了研究这种不同的行为，我使用荧光和圆二色光谱监测了蛋白质随温度变化的结构变化。被丙烯酰胺猝灭的本征色氨酸荧光表明，即使在25℃下，αB-晶体蛋白中的色氨酸比αA-晶体蛋白中的单独色氨酸更容易获得，蛋白质结合的8-苯胺基-1-磺酸荧光表明αB-晶体蛋白疏水表面的溶剂可及性更高。圆二色谱研究表明，αA-晶体蛋白在50℃以上发生了一些三级结构的变化，而αB-晶体蛋白的三级结构发生了显著的变化45℃。我们的研究表明，尽管αB-晶体蛋白具有高度的序列同源性和普遍接受的结构相似性，但它对温度依赖的结构扰动比αA或α-晶体蛋白更敏感，并显示出其伴侣性质的差异。这些差异似乎与α-晶状体蛋白伴侣样活性的温度依赖性增强有关，并表明这两种蛋白质在胁迫条件下作为单独的蛋白质和在α-晶状体蛋白异质聚集体中的不同作用。
alpha-Crystallin, a heteromultimeric protein made up of alpha A- and alpha B-crystallins, functions as a molecular chaperone in preventing the aggregation of proteins. We have shown earlier that structural perturbation of alpha-crystallin can enhance its chaperone-like activity severalfold. The two subunits of alpha-crystallin have extensive sequence homology and individually display chaperone-like activity. We have investigated the chaperone-like activity of alpha A- and alpha B-crystallin homoaggregates against thermal and nonthermal modes of aggregation. me find that, against a nonthermal mode of aggregation, alpha B-crystallin shows significant protective ability even at subphysiological temperatures, at which alpha A-crystallin or heteromultimeric alpha-crystallin exhibit very little chaperone-like activity. Interestingly, differences in the protective ability of these homoaggregates against the thermal aggregation of beta(L)-crystallin is negligible. To investigate this differential behavior, me have monitored the temperature-dependent structural changes in both the proteins using fluorescence and circular dichroism spectroscopy. Intrinsic tryptophan fluorescence quenching by acrylamide shows that the tryptophans in alpha B-crystallin are more accessible than the lone tryptophan in alpha A-crystallin even at 25 degrees C. Protein-bound 8-anilinon-aphthalene-1-sulfonate fluorescence demonstrates the higher solvent accessibility of hydrophobic surfaces on alpha B-crystallin. Circular dichroism studies show some tertiary structural changes in alpha A-crystallin above 50 degrees C, alpha B-crystallin, on the other hand, shows significant alteration of tertiary structure by 45 degrees C, Our study demonstrates that despite a high degree of sequence homology and their generally accepted structural similarity, alpha B-crystallin is much more sensitive to temperature-dependent structural perturbation than alpha A- or alpha-crystallin and shows differences in its chaperone-like properties. These differences appear to be relevant to temperature-dependent enhancement of chaperone-like activity of alpha-crystallin and indicate different roles for the two proteins both in alpha-crystallin heteroaggregate and as separate proteins under stress conditions.