The three-dimensional structure of two mutants of the signal transduction protein CheY suggest its molecular activation mechanism
The three-dimensional structure of two mutants of the signal transduction protein CheY suggest its molecular activation mechanism
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DOI：
10.1006/jmbi.1996.0151
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发表时间：
1996-03-22
影响因子：
5.6
通讯作者：
Coll, M
中科院分区：
文献类型：
--
作者：
Bellsolell, L;Cronet, P;Coll, M
反应调节蛋白CheY的单突变体M17 G和三突变体F14 G-S15 G-M17 G的三维晶体结构已被确定为分别为2.3和1.9埃。如通过稳定性的变化所确定的，两种突变体都结合必需的Mg 2+阳离子，但结合不会引起在野生型蛋白中观察到的W58的内在荧光猝灭。环β 4-α 4在两种突变体中似乎都非常灵活，并且螺旋α 4在天然Mg 2 +-CheY中起始于N94，在天然apo-CheY中起始于K91，在两种突变体中起始于残基K92。由于M17 G突变释放的空间，K109的侧链似乎更移动的。在三重突变体中，K109和相邻残基（环β 5-α 5）的主链被置换几乎2埃，影响残基T87至E89（β 4的C末端）处的主链。三重突变体结构具有在活性位点结合的Mg 2+，但是尽管Mg 2+配位与天然Mg 2 +-CheY的配位相似，但金属结合的结构后果是完全不同的。这些突变似乎破坏了在天然蛋白质中观察到的运动传递机制。我们认为，K109的侧链，在天然蛋白质中的V86，A88和M17之间包装，在激活和失活时向前和向后滑动，在环β 5-α 5处拖动主链，并在蛋白质的功能表面引发更大的运动。(C)1996年学术出版社
The three-dimensional crystal structures of the single mutant M17G and the triple mutant F14G-S15G-M17G of the response regulator protein CheY have been determined to 2.3 and 1.9 Angstrom, respectively Both mutants bind the essential Mg2+ cation as determined by the changes in stability, but binding does not cause the intrinsic fluorescence quenching of W58 observed in the wild-type protein. The loop beta 4-alpha 4 appears to be very flexible in both mutants and helix alpha 4, which starts at N94 in the native Mg2+-CheY and at K91 in the native apo-CheY, starts in both mutants at residue K92. The side-chain of K109 appears to be more mobile because of the space freed by the M17G mutation. In the triple mutant the main chain of K109 and adjacent residues (loop beta 5-alpha 5) is displaced almost by 2 Angstrom affecting the main chain at residues T87 to E89 (C terminus of beta 4). The triple mutant structure has a Mg2+ bound at the active site, but although the Mg2+ coordination is similar to that of the native Mg2+-CheY, the structural consequences of the metal binding are quite different. It seems that the mutations have disrupted the mechanism of movement transmission observed in the native protein. We suggest that the side-chain of K109, packed between V86, A88 and M17 in the native protein, slides forwards and backwards upon activation and deactivation dragging the main chain at the loop beta 5-alpha 5 and triggering larger movements at the functional surface of the protein. (C) 1996 Academic Press Limited