[Effect of growth, temperature and media on the expression of secE promoter of Streptomyces lividans TK24].
Streptomyces are Gram-positive, filamentous soil bacteria, which produce a wide variety of metabolites that are currently being exploited in both medicine and agriculture. Moreover, Streptomyces lividans is used as a host strain to effectively express and secrete heterologous proteins to culture media. Genes encoding Sec proteins responsible for the translocation of preproteins have been identified in S. lividans. SecYEG, a complex of integral membrane proteins SecY, SecE, and SecG in the cytoplasmic membrane, constitutes a pathway for polypeptide movement. SecA plays a central role in translocation as it is the site of preprotein entry into the translocase and it is the only ATPase essential for preprotein translocation. The role of SecD and SecF is to regulate the movement of the translocating protein. A better understanding of their regulatory mechanism could help to develop S. lividans strains with hyper-secretory capacity. In this study, a reporter system was used to investigate the regulatory mechanism of secE promoter. Upstream sequence (496 bp) of secE of S. lividans TK24 was cloned and characterized in a promoter probe vector pIJ4083 upstream of promoterless xylE. Sequencing analysis revealed that secE upstream sequence of S. lividans shares 99.8% homology with the secE promoter of S. coelicolor. The transcriptive activity of this fragment approximates vsi promoter in CMI medium, a strong promoter from S. venezuelae. The activity of secE promoter was observed in different growth phase, culture temperature and culture media. The expression level of secE was higher during the log phase, but decreased at the beginning of the stationary phase. At growth temperature of 28 degrees C, the expression level was much higher than that at 37 degrees C. When the bacteria were incubated in NB, Phage, and CM medium respectively, the expression level of secE promoter was different at a certain fermentation time, but all reached the highest level at log phase. Further analysis revealed that glucose may repress secE promoter expression.
Wang
Li-Fei
LF
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China.
Hong
Bin
B
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects / Q000235:genetics; D003470:Culture Media / Q000494:pharmacology; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D013302:Streptomyces / Q000235:genetics*; D013696:Temperature; D013997:Time Factors
D001426:Bacterial Proteins; D003470:Culture Media; D012333:RNA, Messenger