Lethality of a dut (deoxyuridine triphosphatase) mutation in Escherichia coli.
A chloramphenicol resistance gene was cloned into a plasmid-borne dut gene, producing an insertion mutation that was then transferred to the chromosome by allelic exchange. The mutation could not be acquired by haploid strains through substitutive recombination, even when two flanking markers were simultaneously transduced. The insertion was easily transferred, via generalized transduction, into the chromosomal dut region of strains harboring a lambda dut + transducing phage; however, the resulting dut mutant/lambda dut + merodiploid could not then be cured of the prophage. This apparent lethality of the mutation could not be explained by effects on adjacent genes; the dfp gene retained complementing activity, and a ttk insertion mutant was viable. The dut gene product, deoxyuridine triphosphatase, is known to reduce incorporation of uracil into DNA and to be required in the de novo synthesis of thymidylate. Therefore, an attempt was made to determine whether the dut insertion would be tolerated in strains carrying the following compensatory mutations: dcd (dCTP deaminase) and cdd (deoxycytidine deaminase), which should reduce dUTP formation; ung (uracil-DNA glycosylase), which should reduce fatally excessive excision repair; deoA (thymidine phosphorylase), which should enhance the utilization of exogenous thymidine; and sulA, which should reduce the lethal side effects of SOS regulon induction. These mutations, either alone or in various combinations, did not permit the survival of a haploid dut insertion mutant, suggesting that the dut gene product might have an essential function apart from its deoxyuridine triphosphatase activity.
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Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
Zhang
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Weiss
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D004252:DNA Mutational Analysis; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial; D005804:Genes, Lethal; D009876:Operon; D011755:Pyrophosphatases / Q000235:genetics*; D013696:Temperature; D013936:Thymidine / Q000378:metabolism; D014161:Transduction, Genetic
D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase; D013936:Thymidine