Summary
The DNA polymerase induced by bacteriophage T7 is composed of a phage-specified subunit, the gene 5 protein, and a host-specified subunit, the 12,000 dalton thioredoxin of Escherichia coli. tsnC mutants of E. coli B (Chamberlin, 1974) have no detectable thioredoxin, and thus cannot support the growth of phage T7, although they are killed by phage infection. A mutant of E. coli K12 affecting thioredoxin has been isolated by a modification of the procedure used by Chamberlin (1974) to isolate tsnC mutants of E. coli B. The gene affecting thioredoxin has been designated trxA. This mutant, E. coli JM109, shows the TsnC phenotype in that it is killed by, but cannot support the growth of, bacteriophage T7. T7 DNA replication does not occur in mutantinfected cells. These phenotypic expressions of the tsnC mutation have enabled us to screen recombinants for the trxA allele in HfrxF- crosses and F-ductants in episome transfer experiments. Extracts of transductants in generalized transduction by P1 phage were screened for their ability to complement partially purified phage T7 gene 5 protein to form T7 DNA polymerase. The trxA gene is located at 84 min on the E. coli linkage map, between uvrE and metE; trxA is 34% co-transducible with metE.
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Aldeberg, E.A., Mandel, M., Chen, G.C.C.: Optimal conditions for mutagenesis by N-methyl-N′-nitro-N-nitrosoguanidine in Escherichia coli K12. Biochem. biophys. Res. Commun. 18, 788–795 (1965)
Bachmann, B.J., Low, K.B., Taylor, A.L.: Recalibrated linkage map of Escherichia coli K12. Bact. Rev. 40, 116–167 (1976)
Campbell, J.L., Soll, L., Richardson, C.C.: Isolation and partial characterization of a mutant of Escherichia coli deficient in DNA polymerase II. Proc. nat. Acad. Sci. (Wash.) 69, 2090–2094 (1972)
Chamberlin, M.: Isolation and characterization of prototrophic mutants of Escherichia coli unable to support the intracellular growth of T7. J. Virol. 14, 509–516 (1974)
Fuchs, J.: Isolation of an Escherichia coli mutant deficient in thioredoxin reductase. J. Bact. 129, 967–972 (1977)
Fuchs, J.A., Karlström, H.O.: Mapping of ndrA and ndrB in Escherichia coli K12. J. Bact. 128, 810–814 (1976)
Fuchs, J.A., Karlström, H.O., Warner, H.R., Reichard, P.: Defective gene product in dnaF mutant of Escherichia coli. Nature (Lond.) New Biol. 238, 69–71 (1972)
Fuchs, J.A., Warner, H.R.: Isolation of an Escherichia coli mutant deficient in glutathione synthesis. J. Bact. 124, 140–148 (1975)
Grippo, P., Richardson, C.C.: Deoxyribonucleic acid polymerase of bacteriophage T7. J. biol. Chem. 246, 6867–6873 (1971)
Hinkle, D.C., Richardson, C.C.: Bacteriophage T7 deoxyribonucleic acid replication in vitro. Requirements for deoxyribonucleic acid synthesis and characterization of the product. J. biol. Chem. 249, 2974–2984 (1974)
Holmgren, A.: Hydrogen donor system for Escherichia coli ribonucleoside-diphosphate reductase dependent upon glutathione. Proc. nat. Acad. Sci. (Wash.) 73, 2275–2279 (1976)
Holmgren, A., Söderberg, B.-O., Eklund, H., Bränden, C.-I.: Three-dimensional structure of Escherichia coli thioredoxin-S2 to 2.8 Å resolution. Proc. nat. Acad. Sci. (Wash.) 72, 2305–2309 (1975)
Laurent, T.C., Moore, E.C., Reichard, P.: Enzymatic synthesis of deoxyribonucleotides. IV. Isolation and characterization of thioredoxin, the hydrogen donor from Escherichia coli B. J. biol. Chem. 239, 3436–3444 (1964)
Lennox, E.S.: Transduction of linked genetic character of the host by bacteriophage P1. Virology 1, 190–206 (1955)
Low, B., Wood, T.H.: A quick and efficient method for interruption of bacterial conjugation. Genet. Res. 6, 300–303 (1965)
Low, K.B.: Escherichia coli K12 F-prime factors, old and new. Bact. Rev. 36, 587–607 (1972)
Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J.: Protein measurement with the Folin phenol reagent. J. biol. Chem. 193,265–275 (1951)
Mark, D.F., Richardson, C.C.: Escherichia coli thioredoxin: A subunit of bacteriophage T7 DNA polymerase. Proc. nat. Acad. Sci. (Wash.) 73, 780–784 (1976)
Mead, C.G.: The enzymatic condensation of oligonucleotides with polydeoxyribonucleotides. Proc. nat. Acad. Sci. (Wash.) 52, 1482–1488 (1964)
Milcarek, C., Weiss, B.: Preliminary mapping of mutations affecting exonuclease III in Escherichia coli K-12. J. Bact. 113, 1086–1088 (1973)
Miller, J.H.: Experiments in molecular genetics. Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory 1972
Modrich, P., Richardson, C.C.: Bacteriophage T7 deoxyribonculeic acid replication in vitro. A protein of Escherichia coli required for bacteriophage T7 DNA polymerase activity. J. biol. Chem. 250, 5508–5514 (1975a)
Modrich, P., Richardson, C.C.: Bacteriophage T7 deoxyribonucleic acid replication in vitro. Bacteriophage T7 DNA polymerase: An enzyme of phage- and host-specified subunits. J. biol. Chem. 250, 5515–5522 (1975b)
Porque, P.G., Baldesten, A., Reichard, P.: The involvement of the thioredoxin system in the reduction of methionine sulfoxide and sulfate. J. biol. Chem. 245, 2371–2374 (1970)
Reichard, P.: In: The Biosynthesis of Deoxyribose. Ciba Lectures in Biochemistry. New York: John Wiley and Sons 1967
Rosner, J.L.: Formation, induction, and curing of bacteriophage P1 lysogens. Virology 48, 679–689 (1972)
Siegal, E.C.: Ultraviolet-sensitive mutator strain of Escherichia coli K12. J. Bact. 113, 145–160 (1973)
Studier, F.W.: The genetics and physiology of bacteriophage T7. Virology 39, 562–574 (1969)
Tsang, M., Schiff, J.A.: Sulfate reducing pathway in E. coli involving bound intermediates. J. Bact. 125, 923–933 (1976)
Vogel, M.J., Bonner, D.M.: Acetylornithase of Escherichia coli: Partial purification and some properties. J. biol. Chem. 218, 97–106 (1956)
Weber, K., Osborne, M.: The relaibility of molecular weight determinations by dodecyl sulfate-polyacrylamide gel electrophoresis. J. biol. Chem. 244, 4406–4412 (1969)
Wechsler, J.A., Gross, J.D.: Escherichia coli mutants, temperaturesensitive for DNA synthesis. Molec. gen. Genet. 113, 273–284 (1971)
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Communicated by W. Gilbert
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Mark, D.F., Chase, J.W. & Richardson, C.C. Genetic mapping of trxA, a gene affecting thioredoxin in Escherichia coli K12. Molec. gen. Genet. 155, 145–152 (1977). https://doi.org/10.1007/BF00393153
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DOI: https://doi.org/10.1007/BF00393153