Institut für Industrielle Genetik, Universität Stuttgart, Allmandring 31, 70569, Stuttgart, Germany.
World J Microbiol Biotechnol. 2014 May;30(5):1491-9. doi: 10.1007/s11274-013-1569-9. Epub 2013 Dec 1.
Bacillus pumilus SG2 is a chitinolytic bacterium that produces two chitinases, namely ChiS and ChiL. The chiS and chiL genes are consecutively expressed under a common promoter. Regulation of the chiS and chiL genes is under the control of carbon catabolite repression (CCR) in B. pumilus. This study aimed to investigate the cis-acting elements of the chitinase promoter. For this purpose, we transferred the chiS gene along with its specific promoter to Bacillus subtilis as a host. Primer extension analysis revealed two transcription start sites located 287 and 65 bp upstream of the chiS start codon. The distal promoter was highly compatible with the consensus sequence of the σ(A)-type promoters in B. subtilis, whereas the proximal promoter sequence showed less similarity to the σ(A)-type consensus sequence. A catabolite responsive element (cre), which is required for CCR in Bacillus species, was found to be 136 to 123 bp upstream of the chiS start codon. Interestingly, this cre site was located upstream of the -35 of the proximal promoter and downstream of the distal promoter. Deletion of this cre site sequence rendered the chiS expression constitutive.
短小芽孢杆菌 SG2 是一种产生两种几丁质酶的几丁质分解菌，分别为 ChiS 和 ChiL。chiS 和 chiL 基因在共同的启动子下连续表达。短小芽孢杆菌中的碳分解代谢物阻遏（CCR）控制着 chiS 和 chiL 基因的调控。本研究旨在研究几丁质酶启动子的顺式作用元件。为此，我们将 chiS 基因及其特异性启动子转移到枯草芽孢杆菌中作为宿主。引物延伸分析显示，chiS 起始密码子上游 287 和 65 bp 处有两个转录起始位点。远端启动子与枯草芽孢杆菌中 σ(A)-型启动子的共有序列高度兼容，而近端启动子序列与 σ(A)-型共有序列的相似性较低。发现一个分解代谢物响应元件（cre），它是芽孢杆菌属中 CCR 所必需的，位于 chiS 起始密码子上游 136 到 123 bp 处。有趣的是，这个 cre 位点位于近端启动子的-35 上游和远端启动子的下游。删除该 cre 位点序列会使 chiS 表达组成型。
