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 Bacilus 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 -y-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 BaciUlus subtlis 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.Coultate and Sundaram (5) reported that the molar growth yield (49) of a prototrophic strain of Bacillus stearothermophilus progressively decreases at higher growth temperatures. The molar growth yields of this strain appear to be inversely related to the growth rate and high temperature. At higher temperatures, a larger proportion of the glucose carbon remains incompletely utilized in the medium, mostly as acetate, and energy production is uncoupled from respiration (5). Using another strain of B. stearothermophilus, de Vrij et al. (7) found that the efficiency of energy transduction was decreased at high temperature. Other temperatureinduced alterations of metabolic activities were also reported during growth of other strains of B. stearothermophilus. These include use of alternative catabolic pathways (19) and changes in catabolic repression mechanisms (42), de novo synthesis of enzymes (42), and transport of amino acids (7,42). In addition, cultures of B. stearothermophilus grown at or higher than their optimal temperature for growth generally reached cell densities that were lower than the cell densities of cultures grown under the same conditions at lower temperatures (56, 62). Thus, temperature may induce a reversible shift of metabolic activities in some strains of B. stearothermophilus.We before exponential growth at ...

Cloning and Characterization of the str Operon and Elongation Factor Tu Expression in Bacillus stearothermophilus

“…The ϩ1 nucleotide, determined by primer extension experiments (Fig. 4), was mapped 6 bp downstream of the TAGTCT sequence that has 5 nt identical to the B. stearothermophilus Ϫ10 promoter consensus sequence (TA TTA/CT [52]; Fig. 2).…”

An operon from Lactobacillus helveticus composed of a proline iminopeptidase gene (pepl) and two genes coding for putative members of the ABC transporter family of proteins

“…The highest similarities of the Nterminal half was found with Nod1 (50 YO) from RhiTobitlm legtlminosartlm (Evans & Downie, 1986), MalK (44 YO) from E. coli (Gilson e t al., 1982) and LacK (42 %) from Agrobacteritlm radiobacter (Williams et al, 1992). Instead, the C-terminal half of the 50-7 kDa protein shared the highest similarities with OppF (43 YO) from Lactococctls lactis (Tynkkynen e t al., 1993), GlnQ (40 YO) from Bacilltls stearothermophiltls (Wu & Welker, 1991) and PhnC (37 YO) from E. coli (Makino e t al., 1991). The similarity between the N-and C-terminal parts of the 50.7 kDa protein was 38%.…”

Helicobacter pylori ABC transporter: effect of allelic exchange mutagenesis on urease activity

“…P30750): 187 of 201 residues aligned, 114 were identified as exact matches, 28 conservative replacements were found, and one gap of 3 amino acid residues was introduced (56% amino acid identity and 70% amino acid similarity [identical residues plus conservative replacements]). Other proteins identified include the glutamine transport ATP-binding protein (GlnQ) of Bacillus stearothermophilus (32% amino acid identity and 52% similarity) and the glycine betaine-L-proline transport ATP-binding protein (ProV) of E. coli (30% amino acid identity and 58% similarity over the regions that overlap) (22,59).…”