Department of Microbiology, University of California, One Shields Avenue, Davis, CA 95616-8665, USA.
J Bacteriol. 2010 Feb;192(3):734-45. doi: 10.1128/JB.00172-09. Epub 2009 Dec 4.
In the nitrate-responsive, homodimeric NarX sensor, two cytoplasmic membrane alpha-helices delimit the periplasmic ligand-binding domain. The HAMP domain, a four-helix parallel coiled-coil built from two alpha-helices (HD1 and HD2), immediately follows the second transmembrane helix. Previous computational studies identified a likely coiled-coil-forming alpha-helix, the signaling helix (S helix), in a range of signaling proteins, including eucaryal receptor guanylyl cyclases, but its function remains obscure. In NarX, the HAMP HD2 and S-helix regions overlap and apparently form a continuous coiled-coil marked by a heptad repeat stutter discontinuity at the distal boundary of HD2. Similar composite HD2-S-helix elements are present in other sensors, such as Sln1p from Saccharomyces cerevisiae. We constructed deletions and missense substitutions in the NarX S helix. Most caused constitutive signaling phenotypes. However, strongly impaired induction phenotypes were conferred by heptad deletions within the S-helix conserved core and also by deletions that remove the heptad stutter. The latter observation illuminates a key element of the dynamic bundle hypothesis for signaling across the heptad stutter adjacent to the HAMP domain in methyl-accepting chemotaxis proteins (Q. Zhou, P. Ames, and J. S. Parkinson, Mol. Microbiol. 73:801-814, 2009). Sequence comparisons identified other examples of heptad stutters between a HAMP domain and a contiguous coiled-coil-like heptad repeat sequence in conventional sensors, such as CpxA, EnvZ, PhoQ, and QseC; other S-helix-containing sensors, such as BarA and TorS; and the Neurospora crassa Nik-1 (Os-1) sensor that contains a tandem array of alternating HAMP and HAMP-like elements. Therefore, stutter elements may be broadly important for HAMP function.
在硝酸盐响应的同源二聚体 NarX 传感器中，两个细胞质膜α-螺旋限定了周质配体结合域。HAMP 结构域是由两个α-螺旋（HD1 和 HD2）组成的四螺旋平行卷曲螺旋，紧随第二个跨膜螺旋之后。先前的计算研究在一系列信号蛋白中，包括真核受体鸟苷酸环化酶，鉴定出了一个可能的卷曲螺旋形成的α-螺旋，即信号螺旋（S 螺旋），但其功能仍然不清楚。在 NarX 中，HAMP 的 HD2 和 S 螺旋区域重叠，并且显然形成了一个连续的卷曲螺旋，其在 HD2 的远端边界处具有七肽重复的不连续性。在其他传感器中，如酿酒酵母的 Sln1p，也存在类似的复合 HD2-S 螺旋元件。我们构建了 NarX S 螺旋的缺失和错义突变。大多数突变导致组成型信号表型。然而，在 S 螺旋保守核心内的七肽缺失以及消除七肽不连续性的缺失中，观察到了强烈的诱导表型受损。后一种观察结果阐明了甲基接受趋化蛋白中跨越 HAMP 结构域的七肽不连续性的动态束假说的关键要素（Q. Zhou、P. Ames 和 J. S. Parkinson，Mol. Microbiol. 73:801-814, 2009）。序列比较在传统传感器（如 CpxA、EnvZ、PhoQ 和 QseC）中鉴定了 HAMP 结构域和连续卷曲螺旋样七肽重复序列之间的其他七肽不连续性的例子，以及其他包含 S 螺旋的传感器，如 BarA 和 TorS；以及包含串联交替的 HAMP 和 HAMP 样元件的 Neurospora crassa Nik-1（Os-1）传感器。因此，不连续性元件可能对 HAMP 功能具有广泛的重要性。
