Amino Acid Residues in the GerAB Protein Important in the Function and Assembly of the Alanine Spore Germination Receptor of Bacillus subtilis 168
Amino Acid Residues in the GerAB Protein Important in the Function and Assembly of the Alanine Spore Germination Receptor of Bacillus subtilis 168
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DOI：
10.1128/jb.01397-10
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发表时间：
2011-05-01
影响因子：
3.2
通讯作者：
Moir, Anne
中科院分区：
文献类型：
--
作者：
Cooper, Gareth R.;Moir, Anne
内生孢子萌发需要范例 gerA 操纵子，以响应 L-丙氨酸作为唯一的萌发剂，并且三种蛋白质产物 GerAA、GerAB 和 GerAC 预计将在孢子内膜中形成受体复合物。 GerAB 与单组分转运蛋白的氨基酸-多胺-有机阳离子 (APC) 家族具有同源性，预计是具有 10 个跨膜螺旋的完整膜蛋白。定点突变被引入到 gerAB 基因在染色体上的天然位置。膜跨度内一些带电或潜在螺旋断裂残基的改变会显着影响受体功能。在某些情况下，这可能反映了 GerA 受体复合物的完全丧失，根据起始受体蛋白 GerAC 的缺失来判断，这表明改变的 GerAB 蛋白本身可能不稳定，或者改变的结构使复合物不稳定。具有 L-丙氨酸萌发无效表型但将 GerAC 蛋白保留在接近正常水平的突变体更有可能定义具有功能而非结构重要性的氨基酸残基。 GerAB 和 GerAA 蛋白中的单氨基酸取代可以阻止 GerAC 蛋白掺入孢子中；这提供了强有力的证据，证明特定受体内的蛋白质相互作用，并且这些相互作用是受体组装所必需的。 GerAC 受体亚基的脂蛋白性质也很重要； gerAC1 突变体中前脂蛋白信号序列的氨基酸变化导致孢子中不存在 GerAC 蛋白。
The paradigm gerA operon is required for endospore germination in response to L-alanine as the sole germinant, and the three protein products, GerAA, GerAB, and GerAC are predicted to form a receptor complex in the spore inner membrane. GerAB shows homology to the amino acid-polyamine-organocation (APC) family of single-component transporters and is predicted to be an integral membrane protein with 10 membrane-spanning helices. Site-directed mutations were introduced into the gerAB gene at its natural location on the chromosome. Alterations to some charged or potential helix-breaking residues within membrane spans affected receptor function dramatically. In some cases, this is likely to reflect the complete loss of the GerA receptor complex, as judged by the absence of the germinant receptor protein GerAC, which suggests that the altered GerAB protein itself may be unstable or that the altered structure destabilizes the complex. Mutants that have a null phenotype for L-alanine germination but retain GerAC protein at near-normal levels are more likely to define amino acid residues of functional, rather than structural, importance. Single-amino-acid substitutions in each of the GerAB and GerAA proteins can prevent incorporation of GerAC protein into the spore; this provides strong evidence that the proteins within a specific receptor interact and that these interactions are required for receptor assembly. The lipoprotein nature of the GerAC receptor subunit is also important; an amino acid change in the prelipoprotein signal sequence in the gerAC1 mutant results in the absence of GerAC protein from the spore.