A hydrophilic microenvironment in the substrate-translocating groove of the YidC membrane insertase is essential for enzyme function.
A hydrophilic microenvironment in the substrate-translocating groove of the YidC membrane insertase is essential for enzyme function.
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DOI：
10.1016/j.jbc.2022.101690
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发表时间：
2022-03
期刊：
影响因子：
--
通讯作者：
Dalbey RE
中科院分区：
文献类型：
--
作者：
Chen Y;Sotomayor M;Capponi S;Hariharan B;Sahu ID;Haase M;Lorigan GA;Kuhn A;White SH;Dalbey RE
YidC蛋白家族是膜插入酶，其催化膜蛋白的周质结构域经由位于膜的内小叶内的亲水性凹槽的易位。所有的同源物在这个沟的中心都有一个严格保守的带正电荷的残基。在枯草芽孢杆菌中，已提出带正电荷的残基对于与底物的带负电荷的残基相互作用是必不可少的，支持YidC通过早期静电吸引机制催化插入的假设。在这里，我们提供的数据表明，带正电荷的残基是重要的，而不是它的电荷，但增加的沟槽的亲水性。我们发现，当517位的相邻残基是亲水性或芳香性的时，带正电荷的残基对于大肠杆菌YidC功能是必需的，但是当相邻残基是非极性的时，带正电荷的残基是必需的。此外，溶剂可及性研究支持保守的带正电荷的残基的功能，以保持沟的顶部和中部充分水合的想法。此外，我们证明，大肠杆菌和变形链球菌YidC同系物的功能时，严格保守的精氨酸被替换为带负电荷的残基，提供适当的稳定从相邻的残基。这些组合的结果表明，带正电荷的残基的功能，以保持插入酶活性所必需的沟槽中的亲水性微环境，而不是形成与基板的静电相互作用。
The YidC family of proteins are membrane insertases that catalyze the translocation of the periplasmic domain of membrane proteins via a hydrophilic groove located within the inner leaflet of the membrane. All homologs have a strictly conserved, positively charged residue in the center of this groove. In Bacillus subtilis, the positively charged residue has been proposed to be essential for interacting with negatively charged residues of the substrate, supporting a hypothesis that YidC catalyzes insertion via an early-step electrostatic attraction mechanism. Here, we provide data suggesting that the positively charged residue is important not for its charge but for increasing the hydrophilicity of the groove. We found that the positively charged residue is dispensable for Escherichia coli YidC function when an adjacent residue at position 517 was hydrophilic or aromatic, but was essential when the adjacent residue was apolar. Additionally, solvent accessibility studies support the idea that the conserved positively charged residue functions to keep the top and middle of the groove sufficiently hydrated. Moreover, we demonstrate that both the E. coli and Streptococcus mutans YidC homologs are functional when the strictly conserved arginine is replaced with a negatively charged residue, provided proper stabilization from neighboring residues. These combined results show that the positively charged residue functions to maintain a hydrophilic microenvironment in the groove necessary for the insertase activity, rather than to form electrostatic interactions with the substrates.
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影响因子：
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DOI：
10.1073/pnas.1219486110
发表时间：
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影响因子：
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作者：
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通讯作者：
Schnell, Jason R.
影响因子：
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