caa a22 4500 605506272 CHVBK 20210128100626.0 cr unu---uuuuu 210128e20150601xx s 000 0 eng 10.1007/s00253-015-6437-0 doi (NATIONALLICENCE)springer-10.1007/s00253-015-6437-0 Positive and negative regulation of GlnR in validamycin A biosynthesis by binding to different loci in promoter region [Elektronische Daten] [Shuang Qu, Qianjin Kang, Hang Wu, Lei Wang, Linquan Bai] Validamycin A (VAL-A) is a C7N aminocyclitol antibiotic produced by Streptomyces hygroscopicus var. jinggangensis 5008, which has been widely used as antifungal agent against rice sheath blight disease. VAL-A biosynthesis has been proven to be affected by γ-butyrolactone and temperature. Herein, we showed that GlnR, a global regulator in nitrogen metabolism, is specifically associated with valK-valA intergenic promoter region by DNA-affinity chromatography and MS-based protein identification. Subsequent EMSA and DNase I footprinting assays revealed two GlnR binding sites in this promoter region. Targeted disruption of glnR in S. hygroscopicus 5008 led to a significant increase in the transcription of VAL-A structural genes, albeit the VAL-A production was reduced by 80% and the sporulation of the mutant was impaired. Compared with the wild-type 5008, site-specific mutagenesis of GlnR binding site I enhanced VAL-A production by 2.5-fold, whereas the mutation of GlnR binding site II resulted in a 50% reduction of VAL-A yield. Moreover, tandem mutation of site I in the site II mutant led to a 66% increase of VAL-A production. The result suggested that GlnR not only serves as an inhibitor by binding site I but also as an activator by binding site II for VAL-A biosynthesis. Furthermore, overexpression of glnR in the site I mutant JG45 improved VAL-A production for 41% compared with the control strain containing the vector. Therefore, the obtained data illustrate a novel regulatory feature of the global regulator GlnR. GlnR is firstly proved to act simultaneously as an activator and a repressor in validamycin biosynthesis by binding to different loci within a promoter region of the gene cluster. Springer-Verlag Berlin Heidelberg, 2015 Streptomyces nationallicence GlnR nationallicence Validamycin nationallicence Regulation nationallicence Antibiotics nationallicence Qu Shuang State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut Kang Qianjin State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut Wu Hang State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut Wang Lei State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut Bai Linquan State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/11(2015-06-01), 4771-4783 0175-7598 99:11<4771 2015 99 253 https://doi.org/10.1007/s00253-015-6437-0 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s00253-015-6437-0 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Qu Shuang State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut NATIONALLICENCE 700 1- Kang Qianjin State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut NATIONALLICENCE 700 1- Wu Hang State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut NATIONALLICENCE 700 1- Wang Lei State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut NATIONALLICENCE 700 1- Bai Linquan State Key Laboratory of Microbial Metabolism and School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, 200240, Shanghai, China aut NATIONALLICENCE 773 0- Applied Microbiology and Biotechnology Springer Berlin Heidelberg 99/11(2015-06-01), 4771-4783 0175-7598 99:11<4771 2015 99 253