Thermodynamic and structural compensation in ''size-switch'' core repacking variants of bacteriophage T4 lysozyme
Thermodynamic and structural compensation in ''size-switch'' core repacking variants of bacteriophage T4 lysozyme
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DOI：
10.1006/jmbi.1996.0338
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发表时间：
1996-06-14
影响因子：
5.6
通讯作者：
Matthews, BW
中科院分区：
文献类型：
--
作者：
Baldwin, E;Xu, J;Matthews, BW
先前对噬菌体T4溶菌酶核心内随机产生的多个突变的分析表明，Leu121到Ala(L121A)的“大到小”替换和空间上相邻的“小到大”的Ala129到Met(A129M)的替换可能是相互补偿的。为了验证这一假设，我们产生了单独的变异L121A和A129M，以及双“大小开关”突变体L121A/A129M。为了使交换变得对称，还构建了L121A和A129L的组合，得到L121A/A129L，单个突变都是不稳定的。有些令人惊讶的是，从小到大的替换增加了疏水稳定性，但也会引入菌株，其危害比从大到小的替换要小。Ala129-->；Leu和Ala129-->；Met都抵消了L121a的不稳定约50%。此外，与典型的Leu-->；Ala核心取代相比，Leu-->；Ala使A129L和A129M略有稳定，而Leu-->；Ala核心取代使A129L和A129M不稳定。晶体结构分析表明，侧链和主链的组合调整部分地适应了侧链体积的变化，但程度有限。例如，在L121A/A129L中，由Leu121到Ala的替换所产生的空腔实际上变大了。结果表明，与一个核心残基的体积变化相关的不稳定可以通过相邻残基的补偿性体积变化来具体补偿。然而，完全补偿似乎不太可能，因为很难重建一组同等的相互作用。因此，核心相对于表面残基的演变相对较慢似乎是由于两个因素。首先，单个核心残基的突变导致大小的大幅变化通常会导致稳定性的显著丧失；这种突变可能会被选中。其次，如果埋藏残基中确实发生了体积变化，通常不能通过相邻残基的突变来完全补偿。因此，最有可能的反应往往是回复到母体蛋白质。(C)1996年学术出版社有限公司
Previous analysis of randomly generated multiple mutations within the core of bacteriophage T4 lysozyme suggested that the ''large-to-small'' substitution Leu121 to Ala (L121A) and the spatially adjacent ''small-to-large'' substitution Ala129 to Met (A129M) might be mutually compensating. To test this hypothesis, the individual variants L121A and A129M were generated, as well as the double ''size-switch'' mutant L121A/A129M. To make the interchange symmetrical, the combination of L121A with A129L to give L121A/A129L was also constructed.The single mutations were all destabilizing. Somewhat surprisingly, the small-to-large substitutions, which increase hydrophobic stabilization but can also introduce strain, were less deleterious than the large-to-small replacements. Both Ala129 --> Leu and Ala129 --> Met offset the destabilization of L121A by about 50%. Also, in contrast to typical Leu --> Ala core substitutions, which destabilize by 2 to 5 kcal/mol, Leu121 --> Ala slightly stabilized A129L and A129M. Crystal structure analysis showed that a combination of side-chain and backbone adjustments partially accommodated changes in side-chain volume, but only to a limited degree. For example, the cavity that was created by the Leu121 to Ala replacement actually became larger in L121A/A129L.The results demonstrate that the destabilization associated with a change in volume of one core residue can be specifically compensated by an offsetting volume change in an adjacent residue. It appears, however, that complete compensation is unlikely because it is difficult to reconstitute an equivalent set of interactions. The relatively slow evolution of core relative to surface residues appears, therefore, to be due to two factors. First, a mutation in a single core residue that results in a substantial change in size will normally lead to a significant loss in stability; Such mutations will presumably be selected against. Second, if a change in bulk does occur in a buried residue, it cannot normally be fully compensated by a mutation of an adjacent residue. Thus, the most probable response will tend to be reversion to the parent protein. (C) 1996 Academic Press Limited