RT info:eu-repo/semantics/article
T1 CcaR is an autoregulatory protein that binds to the ccaR and cefD-cmcI promoters of the cephamycin C-clavulanic acid cluster in Streptomyces clavuligerus
A1 Santamarta, Irene
A1 Rodríguez García, Antonio
A1 Pérez Redondo, Rosario
A1 Martín Martín, Juan Francisco
A1 Liras Padín, Paloma
A2 Microbiologia
K1 Biotecnología
K1 Cephamycin C
K1 Clavulanic Acid
K1 Transcriptional regulation
K1 CcaR
K1 Streptomyces clavuligerus
K1 Autorregulation
K1 Biosynthesis gene cluster
K1 Secondary metabolism
K1 2414.02 Fisiología Bacteriana
K1 2415.01 Biología Molecular de Microorganismos
AB [EN] The putative regulatory CcaR protein, which is encoded in the β-lactam supercluster of Streptomyces clavuligerus, has been partially purified by ammonium sulfate precipitation and heparin affinity chromatography. In addition, it was expressed in Escherichia coli, purified as a His-tagged recombinant protein (rCcaR), and used to raise anti-rCcaR antibodies. The partially purified CcaR protein from S. clavuligerus was able to bind DNA fragments containing the promoter regions of the ccaR gene itself and the bidirectional cefD-cmcI promoter region. In contrast, CcaR did not bind to DNA fragments with the promoter regions of other genes of the cephamycin-clavulanic acid supercluster including lat, blp, claR, car-cyp, and the unlinked argR gene. The DNA shifts obtained with CcaR were prevented by anti-rCcaR immunoglobulin G (IgG) antibodies but not by anti-rabbit IgG antibodies. ccaR and the bidirectional cefD-cmcI promoter region were fused to the xylE reporter gene and expressed in Streptomyces lividans and S. clavuligerus. These constructs produced low catechol dioxygenase activity in the absence of CcaR; activity was increased 1.7- to 4.6-fold in cultures expressing CcaR. Amplification of the ccaR promoter region lacking its coding sequence in a high-copy-number plasmid in S. clavuligerus ATCC 27064 resulted in a reduced production of cephamycin C and clavulanic acid, by 12 to 20% and 40 to 60%, respectively, due to titration of the CcaR regulator. These findings confirm that CcaR is a positively acting autoregulatory protein able to bind to its own promoter as well as to the cefD-cmcI bidirectional promoter region
PB American Society for Microbiology
SN 0021-9193
LK https://hdl.handle.net/10612/17714
UL https://hdl.handle.net/10612/17714
NO Santamarta, I., Rodríguez-García, A., Pérez-Redondo, R., Martín, J. F., & Liras, P. (2002). CcaR is an autoregulatory protein that binds to the ccaR and cefD-cmcI promoters of the cephamycin C-clavulanic acid cluster in Streptomyces clavuligerus. Journal of Bacteriology, 184(11), 3106-3113. https://doi.org/10.1128/JB.184.11.3106-3113.2002
DS BULERIA. Repositorio Institucional de la Universidad de León
RD 20-may-2024