Slow solvation dynamics at the active site of an enzyme: Implications for catalysis
Slow solvation dynamics at the active site of an enzyme: Implications for catalysis
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DOI：
10.1021/bi0473915
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发表时间：
2005-06-28
期刊：
影响因子：
2.9
通讯作者：
Bhattacharyya, K
中科院分区：
文献类型：
--
作者：
Guha, S;Sahu, K;Bhattacharyya, K
用荧光探针acrylodan（acrylodan）在半胱氨酸残基C229上特异性地连接到酶的活性位点附近，研究了酶活性位点的溶剂化动力学。皮秒的时间依赖性荧光斯托克斯位移表明缓慢的溶剂化动力学在活性位点的酶，在没有任何基板。当底物（谷氨酰胺或tRNA（Gln））与酶结合时，溶剂化动力学变得更慢。构建了谷氨酰胺结合位点被破坏的突变体Y211 H-GlnRS。在C229处用acrylodan标记的突变体Y211 H-GlnRS表现出显著不同的溶剂弛豫，从而证明缓慢的动力学确实与活性位点相关。催化和特异性的影响进行了讨论。
Solvation dynamics at the active site of an enzyme, glutaminyl-tRNA synthetase (GlnRS), was studied using a fluorescence probe, acrylodan, site-specifically attached at cysteine residue C229, near the active site. The picosecond time-dependent fluorescence Stokes shift indicates slow solvation dynamics at the active site of the enzyme, in the absence of any substrate. The solvation dynamics becomes still slower when the substrate (glutamine or tRNA(Gln)) binds to the enzyme. A mutant Y211H-GlnRS was constructed in which the glutamine binding site is disrupted. The mutant Y211H-GlnRS labeled at C229 with acrylodan exhibited significantly different solvent relaxation, thus demonstrating that the slow dynamics is indeed associated with the active site. Implications for catalysis and specificity have been discussed.