Regulatory mechanism of genes involved in chitin degradation system of bacteria

细菌几丁质降解系统相关基因的调控机制

基本信息

批准号：
16580066
负责人：
TSUJIBO Hiroshi
金额：
$ 2.18万
依托单位：
Osaka University of Pharmaceutical Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16580066/
关键词：
marine bacteria actinomycete chitin chitin degradation chitinase gene regulation

项目摘要

We found that four chitinases (ChiA, ChiB, ChiC, and ChiD), three β-N-acetylglucosaminidases (GlcNAcases A, B, and C), a transglycosylative enzyme (Hex99), a chitin-binding protein (Cbp1), a serine protease (AprIV) and a metalloprotease (MprIII) were involved in chitin degradation system of Pseudoalteromonas sp. strain O-7. AprIV and chitinases were induced in the presence of N-acetylglucosamine (GlcNAc). These results suggest that AprIV and chitinases might be coordinately controlled by the same regulatory system. Recently, we have found that ArcA, which is a response regulator of ArcB/ArcA two-component signal transduction system, binds to the upstream region of aprIV gene. The arcA and arcB genes were cloned and expressed in Escherichia coli. The ArcA protein as a response regulator was transphosphorylated by the ArcB protein as a sensor kinase. The ArcA protein was also phosphorylated by acetylphosphate which is an indicator of nutritive conditions of cells. Transphosphorylation fr … More om ArcB to ArcA was promoted by GlcNAc which is a component of chitin. Generally ArcA-ArcB is a two-component signal transduction system which facilitates adaptation to anoxia. However, our research indicated that ArcA plays an important role even in aerated conditions. Gel retardation assay and BIAcore analysis demonstrated that phosphorylated ArcA (ArcA-P) more tightly bound to the upstream region of aprIV and chiD genes compared to those of chiA, chiB, chiC, and cpb1 genes. Furthermore, footprint analyses indicate that ArcA-P binds to the consensus sequence, 5'-ACAATTAGANNNNNACTAAAACA-3'. The consensus sequence was found near the promoter region of each gene. These results suggest that ArcB/ArcA two-component signal transduction system negatively regulates the expression of genes involved in chitin degradation system of the strain.On the other hand, Streptomyces thermoviolaceus OPC-520 produced four chitinases (Chi40, Chi35, Chi30, Chi25) in the presence of chitin. These genes contain a common similar pair of direct repeat sequences in the promoter regions, which have been suggested to be involved in both chitin induction and glucose repression. We found that regulatory gene involved in induction existed in the upstream region of genes encoding chitobiose transporter. We are doing the cloning and expression of the regulatory gene to examine whether the gene product binds to the direct repeat sequences of chitinase genes. Less

我们发现四种几丁质酶（ChiA、ChiB、ChiC 和 ChiD）、三种 β-N-乙酰氨基葡萄糖苷酶（GlcNAcases A、B 和 C）、一种转糖基酶 (Hex99)、一种几丁质结合蛋白 (Cbp1)、一种丝氨酸蛋白酶（AprIV）和金属蛋白酶（MprIII）参与几丁质降解系统假交替单胞菌菌株 O-7. 和几丁质酶在 N-乙酰氨基葡萄糖 (GlcNAc) 的存在下被诱导。最近，我们发现 ArcA 可能受到相同的调控。是 ArcB/ArcA 双组分信号转导系统的响应调节因子，与 aprIV 基因的上游区域结合。 arcB 基因在大肠杆菌中被克隆并表达，作为反应调节剂的 ArcA 蛋白被作为传感器激酶的 ArcB 蛋白转磷酸化，乙酰磷酸是细胞营养状况的指标。 ArcB 到 ArcA 是由 GlcNAc 促进的，GlcNAc 是几丁质的一个组成部分。一般来说，ArcA-ArcB 是一种有利于适应的双组分信号转导系统。然而，我们的研究表明，即使在通气条件下，ArcA 也发挥着重要作用，BIAcore 分析表明，与 aprIV 和 chiD 基因的上游区域相比，磷酸化 ArcA (ArcA-P) 的结合更紧密。 chiA、chiB、chiC 和 cpb1 基因此外，足迹分析表明 ArcA-P 与共有序列结合。 5'-ACAATTAGANNNNNACTAAAACA-3'在每个基因的启动子区域附近发现了共有序列，这些结果表明ArcB/ArcA双组分信号转导系统负调控参与菌株几丁质降解系统的基因的表达。另一方面，热紫链霉菌 OPC-520 在几丁质存在下产生四种几丁质酶（Chi40、Chi35、Chi30、Chi25）。基因在启动子区域包含一对共同的相似的同向重复序列，这被认为参与几丁质诱导和葡萄糖抑制，我们发现参与诱导的调节基因存在于编码壳二糖转运蛋白的基因的上游区域。进行调控基因的克隆和表达，以检查基因产物是否与几丁质酶基因的直接重复序列结合。