In vitro phosphorylation study of the arc two-component signal transduction system of Escherichia coli
In vitro phosphorylation study of the arc two-component signal transduction system of Escherichia coli
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DOI：
10.1128/jb.179.17.5429-5435.1997
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发表时间：
1997-09-01
影响因子：
3.2
通讯作者：
Lin, ECC
中科院分区：
文献类型：
--
作者：
Georgellis, D;Lynch, AS;Lin, ECC
ArcB和ArcA蛋白构成了一个双组分信号转导系统，在转录调控中起着广泛的作用。在缺氧或环境还原条件下，传感器激酶（ArcB）被刺激以ATP为代价自磷酸化，随后转磷酸化反应调节因子（ArcA）。ArcB是一种复杂的膜结合蛋白，至少包含三个胞质结构域，N-末端具有保守的His292残基的传递结构域（H1），中心具有保守的Asp576残基的接收结构域（D1），和C-末端具有保守的His717残基的交替传递结构域（H2）。我们制备了以下His标记的蛋白质：H1、D1、H2、H1-D1、D1-H2、H1-D1-H2和ArcA。Are蛋白与[gamma-P-32] ATP的各种组合的孵育表明，H1，而不是D1或H2，催化自磷酸化; H1-P将磷酰基转移到D1比转移到ArcA快得多; D1加速H2的转磷酸化。最后，ArcA被H2-P磷酸化的速度比被H1-P磷酸化的速度快得多。可用的数据与信号转导模型一致，其中（i）膜信号的接收触发H1在His292处的自磷酸化，（ii）磷酰基可以迁移到Asp576处的D1，随后迁移到His717处的H2，和（iii）ArcA接受来自His292或His717的磷酰基，其相对贡献由胞质效应物调节。
The ArcB and ArcA proteins constitute a two-component signal transduction system that plays a broad role in transcriptional regulation, Under anoxic or environmentally reducing conditions, the sensor kinase (ArcB) is stimulated to autophosphorylate at the expense of ATP and subsequently transphosphorylates the response regulator (ArcA). ArcB is a complex, membrane-bound protein comprising at least three cytoplasmic domains, an N-terminal transmitter domain with a conserved His292 residue (H1), a central receiver domain with a conserved Asp576 residue (D1), and a C-terminal alternative transmitted-domain,vith a conserved His717 residue (H2), To study the phosphoryl transfer pathways of the Are system, we prepared the following His-tagged proteins: H1, D1, H2, H1-D1, D1-H2, H1-D1-H2, and ArcA. Incubations of various combinations of Are proteins with [gamma-P-32]ATP indicated that H1, but not D1 or H2, catalyzes autophosphorylation; that H1-P transfers the phosphoryl group to D1 much more rapidly than to ArcA; and that D1 accelerates the transphosphorylation of H2. Finally, ArcA is phosphorylated much more rapidly by H2-P than by H1-P, Available data are consistent with a signal transduction model in which (i) reception of a membrane signal(s) triggers autophosphorylation of H1 at His292, (ii) the phosphoryl group can migrate to D1 at Asp576 and subsequently to H2 at His717, and (iii) ArcA receives the phosphoryl group from either His292 or His717, the relative contribution of which is regulated by cytosolic effecters.