Cloning the trpR gene

Abstract

In Escherichia coli, the structural gene for purine nucleoside phosphorylase, deoD, is subject to insertional inactivation by prophage lambda. From one such secondary site lambda lysogen, strain SP265, one may isolate deletions that remove all or part of the trpR gene and other genes in the deo-thr sector of the E. coli chromosome. Specialized transducing phages harboring serB+ and trpR+ were liberated following induction of SP265. All such phages were N-defective, bio-type pseudolysogens whose DNA persisted in the form of plasmids. A collection of transducing phages, differing in their complement of bacterial DNA, was used to locate cleavage sites for BamHI, SalI, and PvuI within the deoD-trpR region of the E. coli genome. The trpR gene lies within a specific 950 base pair BamHI-PvuI segment. A 1250 base pair BamHI fragment carrying a functional trpR gene was cloned into the amplifiable plasmid pBR322. A single SalI site in this fragment was shown to lie within the TrpR gene. In two situations where increased gene dosage might generate elevated amounts of Trp repressor (N-defective trpR+ pseudolysogens and strains harboring pBR322 trpR+ plasmids) neither tryptophan auxotrophy, enhanced sensitivity to DL-5-methyl-tryptophan, nor super repression of the tryptophan biosynthetic enzymes was observed.

arcA (dye), a global regulatory gene in Escherichia coli mediating repression of enzymes in aerobic pathways.

“…The dye gene, whose loss leads to sensitivity to dyes such as toluidine blue, maps near the thr operon (41). We found that the strain ECL547 (arcA ) is dye-resistant, and the mutant ECL549 (arcAl) is dye-sensitive on tryptone/toluidine-blue agar (14).…”

“…Mutations in the dye gene also confer resistance to malespecific phages ( (14). On the basis of altered protein profiles of inner and outer membranes in such mutants, the gene is thought to affect the synthesis of envelope proteins (43,44).…”

Arc and Sfr functions of the Escherichia coli K-12 arcA gene product are genetically and physiologically separable

“…The Escherichia coli gene arcA was identified initially in deletion strains as a locus near trpR conferring sensitivity to redox dyes such as methylene or toluidine blue (Dye phenotype) and resistance of Hfr strains to donor-specific RNA bacteriophages (Msp phenotype) (8,27). Lerner and Zinder (20), Beutin and Achtman (4), and Silverman et al (29) all isolated mutants defective in F-plasmid-dependent DNA donor activity in which the responsible mutation mapped to the trpR-thr region; each laboratory assigned a different mnemonic (fex, sfrA, and cpxC, respectively).…”

Nucleotide sequence and expression of Escherichia coli trpR, the structural gene for the trp aporepressor.

“…aroH codes for one of the three 3-deoxy-2-keto-D-arabinoheptulosonic acid 7-phosphate (DAHP) synthases that catalyze the first step in aromatic amino acid biosynthesis (9,10). trpR was recently cloned from the E. coli chromosome onto phage and plasmid vectors (11,12). The gene is present on a 1.3-kilobase BamHI fragment (11,12).…”