Molecular Cloning and the Nucleotide Sequence of the Clostridium thermocellum trpE Gene1

Abstract

The trpE gene of Clostridium thermocellum, a moderately thermophilic and absolutely anaerobic bacterium, was cloned by its ability to complement growth of an Escherichia coli tryptophan auxotroph (trpE) deficient in anthranilate synthase I. The nucleotide sequence of trpE and its flanking region was determined. The trpE gene overlapped at the termination codon with a putative initiation codon of trpG (trp[G]D), as deduced from the amino acid sequence homology with anthranilate synthase II of Serratia marcescens. S1-nuclease mapping of the trpE transcript produced in E. coli cells suggested that the promoter of C. thermocellum was utilized by E. coli. The amino acid sequence of anthranilate synthase I of C. thermocellum predicted from the nucleotide sequence is more similar to that of an extremely thermophilic bacterium, Thermus thermophilus HB8, than that of mesophilic bacteria.

The Molecular Biology of Tryptophan Synthase: A Model for Protein-Protein Interaction

“…The trp mutant hosts are mostly E. coli derivatives, as used in the cloning of Bacillus stearothermophilus trpBA (Ishiwata et al, 1989), Brevibacterium lactofermentum trpE (Matsui, Miwa and Sano, 1987), Caulobacter crescentus trpF (Winkler et al, 1984), Clostridium thermocellum trpEG (Sato et al, 1989). Lactobacillus casei trpDCFBA (Natori, Kano and Imamoto, 1990), Methanococcus voltae trpFBA (Sibold and Henriquet, 1988), Streptomyces griseus trpCBA (Rivero-Lezcano et al, 1990) and Thermus thermophilus trpEG (Sato et al, 1988).…”

7 The Molecular Biology of Tryptophan Synthase: A Model for Protein–Protein Interaction

“…The trp mutant hosts are mostly E. coli derivatives, as used in the cloning of Bacillus stearothermophilus trpBA (Ishiwata et al, 1989), Brevibacterium lactofermentum trpE (Matsui, Miwa and Sano, 1987), Caulobacter crescentus trpF (Winkler et al, 1984), Clostridium thermocellum trpEG (Sato et al, 1989), Lactobacillus casei trpDCFBA (Natori, Kano and Imamoto, 1990), Methanococcus voltae trpFBA (Sibold and Henriquet, 1988), Streptomyces griseus trpCBA (Rivero-Lezcano et al, 1990) and Thermus thermophilus trpEG (Sato et al, 1988). In cases where problems with an E. coli host were anticipated or encountered, other hosts were used.…”