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. 1991 Sep 25;19(18):4843–4847. doi: 10.1093/nar/19.18.4843

Complementation by detached parts of GGCC-specific DNA methyltransferases.

G Pósfai 1, S C Kim 1, L Szilák 1, A Kovács 1, P Venetianer 1
PMCID: PMC328777  PMID: 1923753

Abstract

Individually inactive N- and C-terminal fragments of the m5C-methyltransferase M.BspRI can complement each other resulting in specific, in vivo methylation of the DNA. This was shown by cloning the coding regions for N- and C-terminal parts of the enzyme in compatible plasmids and co-transforming them into E.coli cells. The enzyme could be detached at several different sites, producing either non-overlapping or partially overlapping fragments capable of complementation. Reconstitution of the active methyltransferase from inactive fragments was demonstrated in vitro, as well. Another GGCC-specific methyltransferase, M.BsuRI, showed a similar complementation phenomenon. Moreover, interspecies complementation was observed between appropriate fragments of the two closely related enzymes M.BspRI and M.BsuRI. Fragments of structurally and functionally more different methyltransferases were unable to complement each other.

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Selected References

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  1. Balganesh T. S., Reiners L., Lauster R., Noyer-Weidner M., Wilke K., Trautner T. A. Construction and use of chimeric SPR/phi 3T DNA methyltransferases in the definition of sequence recognizing enzyme regions. EMBO J. 1987 Nov;6(11):3543–3549. doi: 10.1002/j.1460-2075.1987.tb02681.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bestor T., Laudano A., Mattaliano R., Ingram V. Cloning and sequencing of a cDNA encoding DNA methyltransferase of mouse cells. The carboxyl-terminal domain of the mammalian enzymes is related to bacterial restriction methyltransferases. J Mol Biol. 1988 Oct 20;203(4):971–983. doi: 10.1016/0022-2836(88)90122-2. [DOI] [PubMed] [Google Scholar]
  3. Blumenthal R. M., Gregory S. A., Cooperider J. S. Cloning of a restriction-modification system from Proteus vulgaris and its use in analyzing a methylase-sensitive phenotype in Escherichia coli. J Bacteriol. 1985 Nov;164(2):501–509. doi: 10.1128/jb.164.2.501-509.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  5. Brosius J., Holy A. Regulation of ribosomal RNA promoters with a synthetic lac operator. Proc Natl Acad Sci U S A. 1984 Nov;81(22):6929–6933. doi: 10.1073/pnas.81.22.6929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chang A. C., Cohen S. N. Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol. 1978 Jun;134(3):1141–1156. doi: 10.1128/jb.134.3.1141-1156.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Galakatos N. G., Walsh C. T. Specific proteolysis of native alanine racemases from Salmonella typhimurium: identification of the cleavage site and characterization of the clipped two-domain proteins. Biochemistry. 1987 Dec 15;26(25):8475–8480. doi: 10.1021/bi00399a066. [DOI] [PubMed] [Google Scholar]
  8. Holmgren A., Slabý I. Thioredoxin-C': mechanism of noncovalent complementation and reactions of the refolded complex and the active site containing fragment with thioredoxin reductase. Biochemistry. 1979 Dec 11;18(25):5591–5599. doi: 10.1021/bi00592a011. [DOI] [PubMed] [Google Scholar]
  9. Karreman C., de Waard A. Agmenellum quadruplicatum M.AquI, a novel modification methylase. J Bacteriol. 1990 Jan;172(1):266–272. doi: 10.1128/jb.172.1.266-272.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Karreman C., de Waard A. Cloning and complete nucleotide sequences of the type II restriction-modification genes of Salmonella infantis. J Bacteriol. 1988 Jun;170(6):2527–2532. doi: 10.1128/jb.170.6.2527-2532.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kim P. S., Baldwin R. L. Specific intermediates in the folding reactions of small proteins and the mechanism of protein folding. Annu Rev Biochem. 1982;51:459–489. doi: 10.1146/annurev.bi.51.070182.002331. [DOI] [PubMed] [Google Scholar]
  12. Kim S. C., Pósfai G., Szybalski W. A novel gene-fusing vector: construction of a 5'-GGmCC-specific chimeric methyltransferase, M.BspRI/M.BsuRI. Gene. 1991 Apr;100:45–50. doi: 10.1016/0378-1119(91)90348-f. [DOI] [PubMed] [Google Scholar]
  13. Kiss A., Posfai G., Keller C. C., Venetianer P., Roberts R. J. Nucleotide sequence of the BsuRI restriction-modification system. Nucleic Acids Res. 1985 Sep 25;13(18):6403–6421. doi: 10.1093/nar/13.18.6403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Klimasauskas S., Timinskas A., Menkevicius S., Butkienè D., Butkus V., Janulaitis A. Sequence motifs characteristic of DNA[cytosine-N4]methyltransferases: similarity to adenine and cytosine-C5 DNA-methylases. Nucleic Acids Res. 1989 Dec 11;17(23):9823–9832. doi: 10.1093/nar/17.23.9823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Lauster R., Trautner T. A., Noyer-Weidner M. Cytosine-specific type II DNA methyltransferases. A conserved enzyme core with variable target-recognizing domains. J Mol Biol. 1989 Mar 20;206(2):305–312. doi: 10.1016/0022-2836(89)90480-4. [DOI] [PubMed] [Google Scholar]
  16. Pósfai G., Baldauf F., Erdei S., Pósfai J., Venetianer P., Kiss A. Structure of the gene coding for the sequence-specific DNA-methyltransferase of the B. subtilis phage SPR. Nucleic Acids Res. 1984 Dec 11;12(23):9039–9049. doi: 10.1093/nar/12.23.9039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Pósfai G., Kiss A., Erdei S., Pósfai J., Venetianer P. Structure of the Bacillus sphaericus R modification methylase gene. J Mol Biol. 1983 Nov 5;170(3):597–610. doi: 10.1016/s0022-2836(83)80123-5. [DOI] [PubMed] [Google Scholar]
  18. Pósfai J., Bhagwat A. S., Pósfai G., Roberts R. J. Predictive motifs derived from cytosine methyltransferases. Nucleic Acids Res. 1989 Apr 11;17(7):2421–2435. doi: 10.1093/nar/17.7.2421. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rose R. E. The nucleotide sequence of pACYC184. Nucleic Acids Res. 1988 Jan 11;16(1):355–355. doi: 10.1093/nar/16.1.355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Schimmel P. Hazards and their exploitation in the applications of molecular biology to structure-function relationships. Biochemistry. 1990 Oct 16;29(41):9495–9502. doi: 10.1021/bi00493a001. [DOI] [PubMed] [Google Scholar]
  21. Slabý I., Holmgren A. Structure and enzymatic functions of thioredoxin refolded by complementation of two tryptic peptide fragments. Biochemistry. 1979 Dec 11;18(25):5584–5591. doi: 10.1021/bi00592a010. [DOI] [PubMed] [Google Scholar]
  22. Strittmatter P., Barry R. E., Corcoran D. Tryptic conversion of cytochrome b 5 reductase to an active derivative containing two peptide chains. J Biol Chem. 1972 May 10;247(9):2768–2775. [PubMed] [Google Scholar]
  23. Szilák L., Venetianer P., Kiss A. Cloning and nucleotide sequence of the genes coding for the Sau96I restriction and modification enzymes. Nucleic Acids Res. 1990 Aug 25;18(16):4659–4664. doi: 10.1093/nar/18.16.4659. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Szomolányi E., Kiss A., Venetianer P. Cloning the modification methylase gene of Bacillus sphaericus R in Escherichia coli. Gene. 1980 Aug;10(3):219–225. doi: 10.1016/0378-1119(80)90051-7. [DOI] [PubMed] [Google Scholar]
  25. Taniuchi H., Parr G. R., Juillerat M. A. Complementation in folding and fragment exchange. Methods Enzymol. 1986;131:185–217. doi: 10.1016/0076-6879(86)31042-5. [DOI] [PubMed] [Google Scholar]
  26. Wilson G. G. Organization of restriction-modification systems. Nucleic Acids Res. 1991 May 25;19(10):2539–2566. doi: 10.1093/nar/19.10.2539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Wu J. C., Santi D. V. Kinetic and catalytic mechanism of HhaI methyltransferase. J Biol Chem. 1987 Apr 5;262(10):4778–4786. [PubMed] [Google Scholar]
  28. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  29. Zamenhof P. J., Villarejo M. Construction and properties of Escherichia coli strains exhibiting -complementation of -galactosidase fragments in vivo. J Bacteriol. 1972 Apr;110(1):171–178. doi: 10.1128/jb.110.1.171-178.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]

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