Cloning and characterization of a glutamine transport operon of Bacillus stearothermophilus NUB36: effect of temperature on regulation of transcription

Abstract

We cloned and sequenced a fragment of the Bacillus stearothermophilus NUB36 chromosome that contains two open reading frames (ORFs) whose products were detected only in cells of cultures grown in complex medium at high temperature. The nucleotide sequence of the two ORFs exhibited significant identity to the sequence of the glnQ and glnH loci of the glutamine transport system in enteric bacteria. In addition, growth response to glutamine, sensitivity to the toxic glutamine analog gamma-L-glutamylhydrazide, and glutamine transport assays with parental strain NUB3621 and mutant strain NUB36500, in which the ORF1 coding segment in the chromosome was interrupted with the cat gene, demonstrated that glnQ and glnH encode proteins that are active in the glutamine transport system in B. stearothermophilus. The inferred promoter for the glnQH operon exhibited a low homology to the -35 and -10 regions of the consensus promoter sequences of Bacillus subtilis and Escherichia coli genes. In addition, the inferred promoter for the glnQH operon also exhibited a low homology with the consensus promoter sequence deduced from the sequences of the promoters of nine different genes from B. stearothermophilus. Transcription of the glnQH operon was activated in a nitrogen-rich medium at high temperature and inhibited under the same conditions at low temperature. Transcription of the glnQH operon was partially activated in a nitrogen-poor medium at low temperature. The region upstream from glnQ contains sequences that have a low homology with the nitrogen regulator I-binding sequences and the nitrogen-regulated promoters of enteric bacteria. The effect of temperature on the regulation of the glnQH operon is discussed.