Carbon Catabolite Regulation in Bacillus subtilis

革兰氏阳性菌（如枯草芽孢杆菌）的碳分解代谢调节机制尚不清楚，但已知与大肠杆菌及其亲缘菌中的系统不同。在高水平易于利用的碳源（如葡萄糖）的生长过程中，编码利用次级碳源的酶的基因表达被CcpA抑制，CcpA是一种DNA结合蛋白，可识别靶基因启动子区域的保守操作位点。在这些条件下。CcpA还激活ackA基因的转录，该基因参与以醋酸盐形式排出多余的碳。激活包括将CcpA蛋白结合到与抑制操作位点相同的序列上：对于ackA，目标。ccpa依赖性激活位点位于启动子的上游。CcpA激活ackA转录的分子机制尚不清楚，阐明这一机制是本项目的主要目标。这些研究将包括对激活所需的顺式作用序列的调查，DNA结合研究，以及RNA聚合酶突变效应的分析。该项目的第二个主要目标是鉴定受CcpA转录激活的其他基因。新的靶基因的鉴定将提供CcpA作为枯草芽孢杆菌碳代谢调节剂的全局作用的信息，并将通过分析其保守特征来深入了解转录激活的机制。本项目的第三个主要目标是分析CcpA活性对碳源活性的响应机制。该项目将为微生物代谢控制的分子机制和转录激活机制提供新的信息。枯草芽孢杆菌碳分解代谢调节蛋白CcpA的同源物已经在多种革兰氏阳性生物中被鉴定出来，这表明本文获得的信息可能适用于一些不太直接适用于遗传分析的实验系统。对碳代谢和调控的更深入了解也将提高芽孢杆菌系统在工业生产中的效用。

Henkin 9723091 The mechanisms for carbon catabolite regulation in Gram-positive bacteria such as Bacillus subtilis are not well-understood, but are known to differ from the systems found in Escherichia coli and its relatives. During growth in high levels of a readily utilized carbon source such as glucose, expression of genes encoding enzymes for utilization of secondary carbon sources is repressed by CcpA, a DNA binding protein which recognizes conserved operator sites in the promoter regions of target genes. Under these conditions. CcpA also activates transcription of the ackA gene, which is involved in excretion of excess carbon in the form of acetate. Activation involves binding of CcpA protein to a sequence identical to the operator sites for repression: for ackA, the target. site for CcpA-dependent activation is located upstream of the promoter. The molecular mechanism for transcriptional activation of ackA by CcpA is unknown, and elucidation of this mechanism is a major goal of this project. These studies will involve investigation of cis-acting sequences required for activation, DNA binding studies, and analysis of the effect of RNA polymerase mutations. The second major goal of this project is the identification of additional genes subject to transcriptional activation by CcpA. Characterization of new target genes will provide information about the global role of CcpA as a regulator of carbon metabolism in B. subtilis, and will also provide insight into the mechanism of transcriptional activation through analysis of conserved features. The third major goal of this project is the analysis of the mechanism for the response of CcpA activity to carbon source activity. This project will provide new information about molecular mechanisms for metabolic control in microorganisms, and mechanisms for transcriptional activation. Homologs of the B. subtilis carbon catabolite regulatory protein CcpA have been identified in a variety of Gram-positive organisms, indicating that the information o btained here is likely to apply to a number of experimental systems less directly amenable to genetic analysis. A greater understanding of carbon metabolism and regulation will also improve the utility of Bacillus systems for industrial production.