Ssk1p response regulator binding surface on histidine-containing phosphotransfer protein Ypd1p
Ssk1p response regulator binding surface on histidine-containing phosphotransfer protein Ypd1p
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DOI：
10.1128/ec.2.1.27-33.2003
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发表时间：
2003-02-01
期刊：
影响因子：
--
通讯作者：
West, AH
中科院分区：
文献类型：
--
作者：
Porter, SW;Xu, QP;West, AH
Ypd 1 p是一种含组氨酸的磷酸转移蛋白，在调节酿酒酵母细胞对高渗胁迫反应的His-Asp磷酸转移信号转导通路中起重要作用。Ypd 1 p是从膜结合的Sln 1 p传感器组氨酸激酶到两个下游反应调节蛋白Ssk 1 p和Skn 7 p的磷酰基转移所必需的。为了研究Ypd 1 p与这些反应调节结构域相互作用的分子基础，我们使用了一种方法，该方法将Ypd 1 p中表面暴露的残基的丙氨酸扫描诱变与酵母双杂交相互作用筛选相结合。突变的残基，不利地影响Ypd 1 p与Ssk 1 p的C-末端反应调节结构域的相互作用被确定，并发现集群上或附近的α A螺旋Ypd 1 p。我们的研究结果，通过分析一个模拟的复杂的支持，确定Ypd 1 p的反应调节剂，是由一个保守的疏水残基包围的保守的极性残基簇的结合位点。我们建议，涉及Ypd 1 p的分子相互作用主要是通过疏水接触介导的，而结合特异性和相互作用的强度可能会受到选择极性侧链相互作用的影响。
Ypd1p, a histidine-containing phosphotransfer protein, plays an important role in a branched His-Asp phosphorelay signal transduction pathway that regulates cellular responses to hyperosmotic stress in Saccharomyces cerevisiae. Ypd1p is required for phosphoryl group transfer from the membrane-bound Sln1p sensor histidine kinase to two downstream response regulator proteins, Ssk1p and Skn7p. To investigate the molecular basis for interaction of Ypd1p with these response regulator domains, we used an approach that coupled alanine-scanning mutagenesis of surface-exposed residues in Ypd1p with a yeast two-hybrid interaction screen. Mutated residues that adversely affected the interaction of Ypd1p with the C-terminal response regulator domain of Ssk1p were identified and found to cluster on or near the alphaA helix in Ypd1p. Our results, supported by analysis of a modeled complex, identify a binding site on Ypd1p for response regulators that is composed of a cluster of conserved hydrophobic residues surrounded by less conserved polar residues. We propose that molecular interactions involving Ypd1p are mediated primarily through hydrophobic contacts, whereas binding specificity and strength of interaction may be influenced by select polar side chain interactions.