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 nucleotide sequence of a new insertion sequence (IS) in Escherichiu co/i, IS421, was determined. It is 1340 bp long and contains inverted repeats of 22 bp at its termini. It is flanked by 13 bp direct repeats apparently generated upon insertion. There are two ORFs longer than 200 bp in IS421. One can encode a polypeptide of 371 amino acids (aa) and the other, which is on the other strand, can encode a polypeptide of 102 aa. The C-terminal part of the 371 aa polypeptide shows some homology to that of transposases encoded in some other known IS elements. The copy number of IS421 in chromosomal DNA was 4 for E. coli K-12 and B, and 5 for E. coli C, as determined by the Southern hybridization of restriction fragments.
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