Characterization of the relA/spoT gene from Bacillus stearothermophilus

By use of degenerate primers, we amplified a fragment of a relA/spoT homologous gene from Bacillus stearothermophilus. Chromosomal walking enabled us to sequence the entire gene and its flanking regions. The primary sequence of the gene product is 78% identical to the RelA/SpoT homologue of Bacillus...

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Bibliographic Details
Published in:FEMS microbiology letters 2000-09, Vol.190 (2), p.195-201
Main Authors: Wendrich, Thomas M., Beckering, Carsten L., Marahiel, Mohamed A.
Format: Article
Language:English
Subjects:
Bacillus
Bacillus stearothermophilus
Bacillus subtilis
Bacillus subtilis - genetics
Bacillus subtilis - metabolism
Bacteriology
Biological and medical sciences
chromosome walking
Fundamental and applied biological sciences. Psychology
Gene Deletion
Genes, Bacterial
Genetic Complementation Test
Genetics
Geobacillus stearothermophilus - enzymology
Geobacillus stearothermophilus - genetics
Geobacillus stearothermophilus - growth & development
Guanosine Pentaphosphate - metabolism
Guanosine pentaphosphate synthetase
guanosine-3'-diphosphate-5'-(tri)diphosphate synthase
Guanosine-3′-diphosphate-5′-(tri)diphosphate
Ligases - genetics
Ligases - metabolism
Microbiology
Molecular Sequence Data
Phylogeny
Pyrophosphatases - genetics
Pyrophosphatases - metabolism
RelA
relA gene
Sequence Analysis, DNA
SpoT
spoT gene
Stringent response
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Summary:By use of degenerate primers, we amplified a fragment of a relA/spoT homologous gene from Bacillus stearothermophilus. Chromosomal walking enabled us to sequence the entire gene and its flanking regions. The primary sequence of the gene product is 78% identical to the RelA/SpoT homologue of Bacillus subtilis, and both gene loci share a similar genetic organization. The B. stearothermophilus rel gene was analyzed in vivo by heterologous expression in the B. subtilis relA deletion strain TW30, and is shown to complement the growth defects of TW30. The recombinant Rel Bst protein was detected by Western immunoanalysis, and synthesizes guanosine-3′-diphosphate-5′-(tri)diphosphate ((p)ppGpp) after amino acid stress and carbon starvation. These in vivo data, the genetic organization, and the primary structure compared to other RelA/SpoT homologues provide circumstantial evidence that the identified gene encodes the only (p)ppGpp synthetase in B. stearothermophilus, presumed to serve also as (p)ppGpp hydrolase.
ISSN:0378-1097
1574-6968
DOI:10.1016/S0378-1097(00)00335-9