Open AccessIdentification of a novel mycobacterial transcriptional regulator and its involvement in growth rate dependence and stringent control
Author(s) -
Kamalakannan V,
Ramachandran Geetha,
Narayanan Sujatha,
Vasan S.K.,
Narayanan P.R.
Publication year - 2002
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2002.tb11141.x
Subject(s) - primer extension , biology , microbiology and biotechnology , start codon , mycobacterium smegmatis , gene , transcription (linguistics) , stringent response , lac operon , rna polymerase , promoter , transcriptional regulation , mycobacterium tuberculosis , escherichia coli , gene expression , messenger rna , genetics , medicine , tuberculosis , linguistics , philosophy , pathology
Abstract A novel transcriptional regulator has been identified in the 400‐bp upstream region of the guaA gene of Mycobacterium tuberculosis H37Rv that promotes the expression of lacZ gene in Mycobacterium smegmatis mc 2 155 and M. tuberculosis H37Rv but not in Escherichia coli DH5α. PCR‐mediated deletion mutagenesis and cloning identified a 120‐bp fragment upstream from the guaA gene to be the actual regulator. Primer extension analysis mapped the transcription start site to be the first ‘G’ residue of the translation start codon GTG of the guaA gene. Electrophoretic mobility shift assay showed strong binding of M. smegmatis RNA polymerase holoenzyme to the 400‐bp fragment that expresses lacZ in mycobacterial species and a weak binding to the 280‐bp fragment that expresses only in E. coli DH5α. Both promoter recombinants revealed varied response in the presence of purine nucleotides and exhibited down‐regulation when subjected to amino acid starvation.