Summary
The instability of supercoiled pBR322 DNA obtained from different cells has been investigated. Partially purified plasmid DNA species from rec +, recA and recBC sbcB cells are converted in vitro first to relaxed and then to linear molecules. The recA and recBC sbcB cells produce the best conditions for the monomerization of the pBR322 DNA and the stable maintenance of plasmids. The supercoiled pBR322 DNA from the recBC sbcB recF144 cells has been isolated preferentially in multimeric from (circular oligomers). These DNA forms are not converted to plasmid monomers and are converted to linear molecules three-fold slower than the monomer linearization in the case of the recBC sbcB cells.
On the other hand, incubation of the pure pBR322 DNA with the recF-dependent protein Z (Krivonogov and Novitskaja 1982) results in the ATP-independent conversion of supercoiled plasmid DNA to relaxed and linear molecules. These results demonstrate an endonuclease activity of the recF-controlled protein Z, which may be involved in general recA-dependent recombination and formation of the pBR322 monomers in the cell.
The results also show that the recF144 mutation in recBC sbcB recF and recF cells leads to the absence of detectable amounts of a 49,000 molecular weight protein.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Austin S, Ziese M, Sternberg N (1981) A novel role for site-specific recombination in maintenance of bacterial replicons. Cell 25:729–736
Bresler SE, Krivonogov SV, Lanzov VA (1978) Scale of the genetic map and genetic control of recombination after conjugation in Escherichia coli K-12. Hot spots of recombination. Mol Gen Genet 166:337–346
Bresler SE, Krivonogov SV, Lanzov VA (1979) Genetic determination of the donor properties in Escherichia coli K-12. Phenomena of chromosome mobilization and integrative suppression. Mol Gen Genet 177:177–184
Bresler SE, Krivonogov SV, Lanzov VA (1981) Recombination instability of F′ plasmids in Escherichia coli K-12. Localization of fre-sites. Mol Gen Genet 183:192–196
Busby S, Kolb A, Buc H (1979) Isolation of plasmid-protein complexes from Escherichia coli. Eur J Biochem 99:105–111
Clark AJ (1974) Recombination deficient mutants of Escherichia coli and other bacteria. Annu Rev Genet 7:67–86
Clark AJ (1980) A view of the RecBC and RecF pathways of E. coli recombination. In: Alberts B, Fox CF (eds) Mechanistic studies of DNA replication and genetic recombination, vol-XIX. Academic Press, New York
Clewell DB, Helinski DR (1970) Properties of supercoiled DNA-protein relaxation complex and strand specific of the relaxation event. Biochemistry 9:4428–4440
Eberle H, Van Knapp, Hrynyszyn J, Forrest N (1982) Regulation of DNA synthesis and capacity for initiation in DNA temperature mutants of Escherichia coli. III. Synthesis of the dnaA protein and of DNA-binding proteins. Mol Gen Genet 186:71–77
Ganesan AK, Seawell PS (1975) The effect of lexA and recF mutations on post-replication repair and DNA synthesis in Escherichia coli K-12. Mol Gen Genet 141:189–195
Hakkaart MJJ, Veltkamp E, Nijkamp HJJ (1982) Maintenance of the bacteriocinogenic plasmid CloDF13 in Escherichia coli cells. II. Specific recombination functions involved in plasmid maintenance. Mol Gen Genet 188:338–344
Hanaoka F, Shaw JL, Mueller GC (1979) Recovery of functional proteins from sodium dodecyl sulfate-polyacrylamide gels. Anal Biochem 99:170–174
Hansen FG, Meyenburg K (1979) Characterization of the dnaA, gyrB and other genes in the dnaA region of the Escherichia coli chromosome on specialized transducing phase λ tna. Mol Gen Genet 175:135–144
Holmes DS, Quigley M (1981) A rapid boiling method for the preparation of bacterial plasmids. Anal Biochem 114:193–197
James AA, Morrison PT, Kolodner R (1982) Genetic recombination of bacterial plasmid DNA. Analysis of the effect of recombination-deficient mutations on plasmid recombination. J Mol Biol 160:411–430
Krivonogov SV, Novitskaja VA (1982) A protein, connected with the integrity of the recF gene in Escherichia coli K-12. Mol Gen Genet 187:302–304
Kuwabara N, Uchida M (1981) Functional cooperation of the dnaE and dnaN gene products in Escherichia coli. Proc Natl Acad Sci USA 78:5764–5767
Lilley DMJ (1980) The inverted repeat as a recognizable structural feature in supercoiled DNA molecules. Proc Natl Acad Sci USA 77:6468–6472
Lovett MA, Helinski DR (1975) Relaxation complexes of plasmid DNA and protein. II. Characterization of the proteins associated with the unrelaxed and relaxed complexes of plasmid ColE1. J Biol Chem 250:8790–8795
Meyers JA, Sancher D, Elwell LP, Falkow S (1976) Simple agarose gel electrophoretic method for the identification and characterization of plasmid deoxyribonucleic acid. J Bacteriol 127:1529–1537
Murakami A, Inokuchi H, Hirota Y, Ozeki H, Yamagishi H (1980) Characterization of the dnaA gene carried by lambda transducing phage. Mol Gen Genet 180:235–248
Panayotatos N, Wells RD (1981) Cruciform structures in supercoiled DNA. Nature 289:466–470
Sakakibara Y, Tsukano H, Sako T (1981) Organization and transcription of the dnaA and dnaN genes of Escherichia coli. Gene 13:47–55
Stahl FW (1979) Special sites in generalized recombination. Annu Rev Genet 13:7–24
Yuasa S, Sakakibara Y (1980) Identification of the dnaA and dnaN gene products of Escherichia coli. Mol Gen Genet 180:267–273
Author information
Authors and Affiliations
Additional information
Communicated by H. Böhme
Rights and permissions
About this article
Cite this article
Krivonogov, S.V. The recF-dependent endonuclease from Escherichia coli K12. Formation and resolution of pBR322 DNA multimers. Mol Gen Genet 196, 105–109 (1984). https://doi.org/10.1007/BF00334100
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00334100