The Residue Composition of the Aromatic Anchor of the Second Transmembrane Helix Determines the Signaling Properties of the Aspartate/Maltose Chemoreceptor Tar of Escherichia coli
The Residue Composition of the Aromatic Anchor of the Second Transmembrane Helix Determines the Signaling Properties of the Aspartate/Maltose Chemoreceptor Tar of Escherichia coli
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DOI：
10.1021/bi201555x
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发表时间：
2012-03-06
期刊：
影响因子：
2.9
通讯作者：
Manson, Michael D.
中科院分区：
文献类型：
--
作者：
Adase, Christopher A.;Draheim, Roger R.;Manson, Michael D.
在第二个跨膜螺旋（TM 2）的胞质末端重新定位串联芳香族残基（Trp-209和Tyr-210）调节大肠杆菌（Tar（Ec））的天冬氨酸/麦芽糖化学受体的信号输出。在此，我们通过研究在位置209和210处具有Ala、Phe、Tyr和Trp的所有可能组合的Tar（Ec）变体文库的功能，直接评估了芳香族锚的残基组成的影响。我们确定了芳香锚的三个重要性质。首先，在位置209处的Trp残基需要在不存在适应性甲基化的情况下维持顺时针（CW）信号输出，但是适应性甲基化恢复了所有突变受体产生CW旋转的能力。第二，当芳香族锚被串联Ala残基取代时，信号传导比Ala残基占据位置209和芳香族残基占据位置210时更少受损。最后，当色氨酸是：尽管210位上的一些取代显著影响麦芽糖的趋化性，但209位上的残基的同一性对基线信号输出或天冬氨酸趋化性几乎没有影响。我们构建的所有突变体受体在半固体琼脂游泳板中支持一定水平的天冬氨酸和麦芽糖趋化，但那些在位置209处没有Trp的突变体受体在其基线信号状态下过度甲基化。这些结果显示了TM 2的细胞质芳香锚在维持基线Tar（Ec）信号输出和对引诱剂信号传导的响应性中的重要性。
Repositioning of the tandem aromatic residues (Trp-209 and Tyr-210) at the cytoplasmic end of the second transmembrane helix (TM2) modulates the signal output of the aspartate/maltose chemoreceptor of Escherichia coli (Tar(Ec)). Here, we directly assessed the effect of the residue composition of the aromatic anchor by studying the function of a library of Tar(Ec) variants that possess all possible combinations of Ala, Phe, Tyr, and Trp at positions 209 and 210. We identified three important properties of the aromatic anchor. First, a Trp residue at position 209 was required to maintain clockwise (CW) signal output in the absence of adaptive methylation, but adaptive methylation restored the ability of all of the mutant receptors to generate CW rotation. Second, when the aromatic anchor was replaced with tandem Ala residues, signaling was less compromised than when an Ala residue occupied position 209 and an aromatic residue occupied position 210. Finally, when Trp was: present at position 209, the identity of the residue at position 210 had little effect on baseline signal output or aspartate chemotaxis, although maltose taxis was significantly affected by some substitutions at position 210. All of the mutant receptors we constructed supported some level of aspartate and maltose taxis in semisolid agar swim plates, but those without Trp at position 209 were overmethylated in their baseline signaling state. These results show the importance of the cytoplasmic aromatic anchor of TM2 in maintaining the baseline Tar(Ec) signal output and responsiveness to attractant signaling.