文献类型：期刊文献

英文题名：Spo0A can efficiently enhance the expression of the alkaline protease gene aprE in Bacillus licheniformis by specifically binding to its regulatory region

作者：Zhou, Cuixia[1,2]; Zhou, Huiying[1]; Fang, Honglei[1]; Ji, Yizhi[3]; Wang, Hongbin[1]; Liu, Fufeng[1]; Zhang, Huitu[1]; Lu, Fuping[1]

第一作者：Zhou, Cuixia

机构：[1] Key laboratory of industrial fermentation microbiology, ministry of education, College of biotechnology, Tianjin University of Science &Technology, Tianjin, 300450, China; [2] School of biology and brewing engineering, Taishan University, Taian, 271018, China; [3] Beijing Key Laboratory of Biomass Waste Resource Utilization, College of Biochemistry and Engineering, Beijing Union University, Beijing, 100023, China

第一机构：Key laboratory of industrial fermentation microbiology, ministry of education, College of biotechnology, Tianjin University of Science &Technology, Tianjin, 300450, China

年份：2020

卷号：159

起止页码：444-454

外文期刊名：International Journal of Biological Macromolecules

收录：EI(收录号：20242616439392);Scopus(收录号：2-s2.0-85085210276)

语种：英文

外文关键词：Bacteriology - DNA - DNA sequences - Electrophoretic mobility - Proteins - Transcription

摘要：The expression of enzymes in Bacillus licheniformis, such as the valuable extracellular alkaline protease AprE, is highly regulated by a complex transcriptional regulation mechanism. Here, we found that the transcript abundance of aprE varies >343-fold in response to the supply of nutrients or to environmental challenges. To identify the underlying regulatory mechanism, the core promoter of aprE and several important upstream regulatory regions outside the promoter were firstly confirmed by 5′-RACE and mutagenesis experiments. The specific proteins that bind to the identified sequences were subsequently captured by DNA pull-down experiments, which yielded the transcriptional factors (TFs) Spo0A, CggR, FruR, YhcZ, as well as fragments of functionally unassigned proteins. Further electrophoretic mobility shift assay (EMSA) and DNase I foot-printing experiments indicated that Spo0A can directly bind to the region from ?92 to ?118 nucleotides upstream of the transcription start site, and the deletion of this specific region drastically decreased the production of AprE. Taken together, these results indicated that the expression of aprE was mainly regulated by the interplay between Spo0A and its cognate DNA sequence, which was successfully applied to overproduce AprE in a genetically modified host harboring three aprE expression cassettes. The DNA binding proteins may serve to increase the efficiency of transcription by creating an additional binding site for RNA polymerase. The discovery of this mechanism significantly increases our understanding of the aprE transcription mechanism, which is of great importance for AprE overproduction. ? 2020 Elsevier B.V.