IN-VIVO CHARACTERIZATION OF THE UNORTHODOX BVGS 2-COMPONENT SENSOR PROTEIN OF BORDETELLA-PERTUSSIS
IN-VIVO CHARACTERIZATION OF THE UNORTHODOX BVGS 2-COMPONENT SENSOR PROTEIN OF BORDETELLA-PERTUSSIS
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DOI：
10.1006/jmbi.1995.0245
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发表时间：
1995-05-05
影响因子：
5.6
通讯作者：
GROSS, R
中科院分区：
文献类型：
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作者：
BEIER, D;SCHWARZ, B;GROSS, R
双组分传感器蛋白通常由氨基酸传感器和羧基末端递质结构域组成，所述结构域含有催化组氨酸残基的自磷酸化的激酶活性。在第二步中，将磷酸转移到位于第二组分（响应调节剂）的接收器结构域中的天冬氨酸残基。一些传感器蛋白如百日咳博德特氏菌的BvgS蛋白具有更复杂的结构。BvgS具有额外的C端域，包括通常只在响应调节器中才能找到的接收器和输出模块。通过体内突变和互补分析研究这些BvgS结构域的功能。构建缺乏C-末端结构域或在保守氨基酸中含有突变的BvgS衍生物。所有突变均导致BvgS失活，如通过在B中整合突变等位基因后在转录和翻译水平上毒力因子的表达所测量的。百日咳染色体然而，这些突变体中的一些可以通过反式的各种C-末端BvgS结构域的单独表达来补充野生型表型，这表明截短的和完整的BvgS蛋白的直接相互作用。因此，使用基于λ阻遏物的二聚化探针系统分析细胞质BvgS结构域的二聚化能力。这些结果表明，BvgS有两个二聚化区域，一个在发射器和第二个在C-末端接收器/输出域。此外，几个BvgS杂合蛋白的构建，其中包含的BvgS接收器和输出结构域与类似的结构域的双组分响应调节剂和传感器蛋白EvgS的取代。发现接收器结构域不携带BvgS特异性功能，并且可以被异源接收器结构域交换。然而，BvgS输出结构域不能被替换为输出结构域的相关蛋白质没有失活的BvgS。
Two-component sensor proteins are typically composed of an amino-acid sensory and a carboxy-terminal transmitter domain containing a kinase activity which catalyses the autophosphorylation of a histidine residue. In a second step, the phosphate is transferred to aspartic acid residues located in the receiver domain of the second component, the response regulator. A few sensor proteins such as the BvgS protein of Bordetella pertussis have a more complex structure. BvgS possesses additional C-terminal domains, including receiver and output modules usually found only in the response regulators. The function of these BvgS domains was investigated by mutation and complementation analysis in vivo. BvgS derivatives were constructed lacking the C-terminal domains or containing mutations in conserved amino acids. All mutations caused the inactivation of BvgS as measured by the expression of virulence factors at the transcriptional and translational level after integration of the mutated alleles in the B. pertussis chromosome. However, some of these mutants could be complemented to the wild-type phenotype by the separate expression of various C-terminal BvgS domains in trans indicating a direct interaction of the truncated and complete BvgS proteins. Therefore, the dimerization capacity of the cytoplasmic BvgS domains was analysed using a lambda repressor based dimerization probe system. These results indicated that BvgS has two dimerization regions, one in the transmitter and a second in the C-terminal receiver/output domains. Furthermore, several BvgS hybrid proteins were constructed which contained substitutions of the BvgS receiver and output domains with similar domains of two-component response regulators and of the sensor protein EvgS. It was found that the receiver domain does not carry BvgS-specific functions and can be exchanged by a heterologous receiver domain. However, the BvgS output domain could not be substituted with output domains of the related proteins without inactivation of BvgS.