Skip to main content
Log in

Bacteriophage lambda initiators: Preparation from a strain that overproduces the O and P proteins

  • Published:
Molecular and General Genetics MGG Aims and scope Submit manuscript

Summary

A recombinant plasmid was constructed which carries bacteriophage λ initiator genes O and P under control of tandemly arranged p L and p R promoters. These promoters were repressed by a thermosensitive repressor, cI857, at low temperature, but became active when the culture was incubated at 42° C. Upon elevation of the temperature, the O and P proteins were overproduced to the extent that they constituted several per cent of the total E. coli cellular proteins. Both the O and P proteins have been purified to apparent homogeneity, and were shown to consist of 298 and 233 amino acid residues, respectively. The amino acid composition and the terminal partial amino acid sequence of each protein were determined. Through these analyses, the locations of the O and P genes in the known λ DNA sequence were determined. The termination codon for the O gene overlaps with the initiation codon for the P gene. The purified O protein binds specifically to the replication origin of λ (λ ori) in accordance with our previous observations. The purified P protein inhibits an ATPase activity of dnaB protein.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  • Backman K, Ptashne M, Gilber W (1976) Construction of plasmids carrying the cI gene of bacteriophage λ. Proc Natl Acad Sci USA 73:4174–4178

    Google Scholar 

  • Bernard HU, Remant E, Hershfield, MU, Das HK, Helinski DR, Yanofsky C, Franklin N (1979) Construction of plasmid cloning vehicles that promote gene expression from bacteriophage lambda pL promoter. Gene 5:59–76

    Google Scholar 

  • Bolivar F, Rodriguez RL, Green PJ, Betlach MC, Heyneker HL, Boyer HW, Grosa JH, Falkow S (1977) Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene 2:95–113

    Article  Google Scholar 

  • Chow LT, Davidson N, Berg D (1974) Electron microscope study of the structure of λ dv DNAs. J Mol Biol 86:69–89

    Google Scholar 

  • Clowes RC, Hayes W (1968) Experiments in microbial genetics. John Wiley and Sons Inc, New York, p 187

    Google Scholar 

  • Denniston-Thompson K, Moore DD, Kruger KE, Furth ME, Blattner FR (1977) Physical structure of the replication origin of bacteriophage lambda. Science 198:1051–1056

    Google Scholar 

  • Furth ME, McLeester C, Dove WF (1978a) Specificity determinants for bacteriophage lambda DNA replication I. A chain of interactions that controls the initiation of replication. J Mol Biol 126:195–225

    Google Scholar 

  • Furth ME, Yates JL (1978) Specificity determinants for bacteriophage lambda DNA replication II. Structure of O protein of λ-ϕ80 and λ-82 hybrid phages and of a λ mutant defective in the origin of replication. J Mol Biol 126:227–240

    Google Scholar 

  • Furth ME, Yates JL, Dove WF (1978b) Positive and negative control of bacteriophage lambda DNA replication. Cold Spring Harbor Symp Quant Biol 43:147–153

    Google Scholar 

  • Goldberg AR, Howe M (1969) New mutations in the S cistron of bacteriophage lambda affecting host cell lysis. Virology 38:200–201

    Google Scholar 

  • Gottesman S, Gottesman M, Shaw JE, Pearson ML (1981) Protein degradation in E.coli: The lon mutation and bacteriophage lambda N and cII protein stability. Cell 24:225–233

    PubMed  Google Scholar 

  • Goulian M, Lucas ZJ, Kornberg A (1968) Enzymatic synthesis of deoxyribonucleic acid XXV. Purification and properties of deoxyribonucleic acid polymerase induced by infection with phage. T4. J Biol Chem 243:627–638

    Google Scholar 

  • Grosschedl R, Hobom G (1979) DNA sequence and structural homologies of the replication origins of lambdoid bacteriophages. Nature 277:621–627

    Google Scholar 

  • Haggerty DM, Schleif RF (1976) Location on bacteriophage lambda DNA of cleavage sites of the site-specific endonuclease from Bacillus amyloliquefaciens H1. J Virol 18:659–663

    Google Scholar 

  • Jeppson J-O, Sjöquist J (1967) Thin-layer chromatography of PTH (Phenylthiohydantoin) amino acid. Anal Biochem 18:264–269

    Google Scholar 

  • Klein A, Bremer B, Kluding H, Symmons P (1978) Initiation of lambda DNA replication in vitro promoted by isolated P-gene product. Eur J Biochem 83:59–66

    Google Scholar 

  • Klinkert J, Klein A (1979) Cloning of the replication gene P of bacteriophage lambda: Effects of increased P−protein synthesis on cellular and phage DNA replication. Mol Gen Genet 171:219–227

    Google Scholar 

  • Kuypers B, Reiser W, Klein A (1980) Cloning of the replication gene O of E. coli bacteriophage lambda and its expression under the control of the lac promoter. Gene 10:195–203

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature (London) New Biol 227:680–685

    Google Scholar 

  • Lipińska B, Podhajska A, Taylor K (1980) Synthesis and decay of λ DNA replication proteins in minicells. Biochem Biophys Res Commun 92:120–126

    Google Scholar 

  • Matsubara K (1974) Preparation of plasmid λ dv from bacteriophage λ: Role of promoter-operator in the plasmid replicon. J Virol 13:596–602

    Google Scholar 

  • Matsubara K (1981) Replication control system in lambda dv. Plasmid 5:32–52

    Google Scholar 

  • Maxam G, Gilbert W (1977) A new method for sequencing DNA. Proc Natl Acad Sci USA 74:560–564

    PubMed  Google Scholar 

  • Murotsu T, Tanaka H, Imaji M, Koga H, Matsubara K, Horiuchi T (1977) Purification and properties of a λ operator-binding protein which is expected to be autorepressor (tof protein) from E. coli carrying λ dv plasmid. Mol Gen Genet 157:139–147

    Google Scholar 

  • Ogawa T, Tomizawa J (1968) Replication of bacteriophage DNA I. Replication of DNA of lambda phage defective in early functions. J Mol Biol 38:217–225

    PubMed  Google Scholar 

  • Remant E, Stanssens P, Fiers W (1981) Plasmid vectors for high-efficient expression controlled by the pL promoter of coliphage lambda. Gene 15:81–93

    Google Scholar 

  • Rosenberg M, Court D, Shimatake H, Brady C, Wulff D (1978) The relationship between function and DNA sequence in an intercistronic regulatory region in phage λ. Nature 272:419–423

    Google Scholar 

  • Scherer G (1978) Nucleotide sequence of the O gene and of the origin of replication in bacteriophage lambda DNA. Nucleic Acids Res 5:3141–3156

    Google Scholar 

  • Schwarz E, Scherer G, Hobom G, Kössel H (1980) The primary structure of the phage λ P gene completes the nucleotide sequence of the plasmid λdvh93. Biochemistry international 1:386–394

    Google Scholar 

  • Shimatake H, Rosenberg M (1981) Purified λ regulatory protein cII positively activates promoters for lysogenic development. Nature 292:128–132

    Google Scholar 

  • Skalka AM (1977) DNA replication-bacteriophage lambda. Curr Top Microbiol Immunol 78:201–237

    Google Scholar 

  • Tomizawa J-I (1971) Functional cooperation of genes O and P. In Hershey AD (ed) The bacteriophage lambda. Cold Spring Harbor Laboratory, New York, pp 549–552

    Google Scholar 

  • Tsurimoto T, Matsubara K (1981a) Purification of bacteriophage λ O protein that specifically binds to the origin of replication. Mol Gen Genet 181:325–331

    Google Scholar 

  • Tsurimoto T, Matsubara K (1981b) Purified bacteriophage λ O protein binds to four repeating sequences at the λ replication origin. Nucleic Acids Res 9:1789–1799

    Google Scholar 

  • Wickner S, Hurwitz J (1975) Interaction of Escherichia coli dnaB and dnaC(D) gene products in vtiro. Proc Natl Acad Sci USA 72:921–925

    Google Scholar 

  • Wickner SH (1978) DNA replication proteins of Escherichia coli and phage λ. Cold Spring Harbor Symp Quant Biol 43:303–310

    Google Scholar 

  • Yates JL, Gette WR, Furth ME, Nomura M (1977) Effects of ribosomal mutations on the read-through of a chain termination signal: Studies on the synthesis of bacteriophage λ O gene protein in vitro. Proc Natl Acad Sci USA 74:689–693

    Google Scholar 

  • Zimmerman CL, Appella E, Pisano JJ (1976) Rapid analysis of amino acid phenythiohydantoins by high-performance liqid chromatography. Anal Biochem 77:569–573

    Google Scholar 

  • Zylicz M, Taylor K (1981) Interactions between phage λ replication proteins, λ DNA and minicell membrane. Eur J Biol 113:303–309

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Additional information

Communicated by T. Yura

Rights and permissions

Reprints and permissions

About this article

Cite this article

Tsurimoto, T., Hase, T., Matsubara, H. et al. Bacteriophage lambda initiators: Preparation from a strain that overproduces the O and P proteins. Molec. Gen. Genet. 187, 79–86 (1982). https://doi.org/10.1007/BF00384387

Download citation

  • Received:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00384387

Keywords

Navigation