SanG, a transcriptional activator, controls nikkomycin biosynthesis through binding to the sanN-sanO intergenic region in Streptomyces ansochromogenes.
Streptomyces ansochromogenes SanG is a pathway-specific regulator that mainly controls the transcription of two transcriptional units involved in nikkomycin biosynthesis. SanG consists of three major functional domains: an N-terminal Streptomyces antibiotic regulatory protein (SARP) domain, a central ATPase domain, and a C-terminal half homologous to guanylate cyclases belonging to the LuxR family. SanG was expressed in Escherichia coli as a C-terminally His(6)-tagged protein. The purified SanG-His( 6) was shown to be a dimer in solution by dynamic light scattering. An electrophoretic mobility-shift assay showed that the purified SanG protein could bind to the DNA fragment containing the bidirectional sanN-sanO promoter region. The SanG-binding sites within the bidirectional sanN-sanO promoter region were determined by footprinting analysis and identified a consensus-directed repeat sequence 5'-CGGCAAG-3'. SanG showed significant ATPase/GTPase activity in vitro, and addition of ATP/GTP enhanced the affinity of SanG for target DNA, but ATP/GTP hydrolysis was not essential for SanG binding to the target DNA. However, real-time reverse transcription PCR showed that mutation of the ATPase/GTPase domain of SanG significantly decreased the transcriptional level of sanN-I and sanO-V. These results indicated that the ATPase/GTPase activity of SanG modulated the transcriptional activation of SanG target genes during nikkomycin biosynthesis.
He
Xihong
X
Graduate School of Chinese Academy of Sciences, Beijing100039, China.|The Key Laboratory of Systematic Mycology and Lichenology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Li
Rui
R
Graduate School of Chinese Academy of Sciences, Beijing100039, China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Pan
Yuanyuan
Y
The Key Laboratory of Systematic Mycology and Lichenology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Liu
Gang
G
The Key Laboratory of Systematic Mycology and Lichenology, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.|State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
Tan
Huarong
H
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000617:Aminoglycosides / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D018983:DNA Footprinting; D021901:DNA, Intergenic / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D020558:GTP Phosphohydrolases / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006868:Hydrolysis; D016297:Mutagenesis, Site-Directed; D013302:Streptomyces / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation
D000617:Aminoglycosides; D001426:Bacterial Proteins; D021901:DNA, Intergenic; C011952:nikkomycin; D000251:Adenosine Triphosphatases; D020558:GTP Phosphohydrolases