Abstract
The time course of the EcoRI endonuclease catalysed cleavage of three substrates, two plasmid DNAs and one oligonucleotide, each with two EcoRI sites, was measured. The two plasmid DNAs with the EcoRI sites 318 and 96 base pairs apart are cut in a distributive fashion, while the oligonucleotide with the EcoRI sites 8 base pairs apart is cut in a partially processive manner. It is concluded that a linear diffusion of the EcoRI endonuclease on its substrate across long stretches of DNA is not likely to be operative during the recognition process. Microscopic dissociation-reassociation processes, however, increase the probability of the enzyme to attack further sites located in the immediate vicinity of a given site.
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Selected References
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- Berg O. G., Winter R. B., von Hippel P. H. Diffusion-driven mechanisms of protein translocation on nucleic acids. 1. Models and theory. Biochemistry. 1981 Nov 24;20(24):6929–6948. doi: 10.1021/bi00527a028. [DOI] [PubMed] [Google Scholar]
- Domdey H., Gross H. J. Gradient thin-layer chromatography of oligonucleotides on DEAE-cellulose: an alternative to homochromatography. Anal Biochem. 1979 Oct 1;98(2):346–352. doi: 10.1016/0003-2697(79)90152-0. [DOI] [PubMed] [Google Scholar]
- Goppelt M., Pingoud A., Maass G., Mayer H., Köster H., Frank R. The interaction of the EcoRI restriction endonuclease with its substrate. A physico-chemical study employing natural and synthetic oligonucleotides and polynucleotides. Eur J Biochem. 1980 Feb;104(1):101–107. doi: 10.1111/j.1432-1033.1980.tb04405.x. [DOI] [PubMed] [Google Scholar]
- Halford S. E., Johnson N. P., Grinsted J. The EcoRI restriction endonuclease with bacteriophage lambda DNA. Kinetic studies. Biochem J. 1980 Nov 1;191(2):581–592. doi: 10.1042/bj1910581. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hillen W., Klein R. D., Wells R. D. Preparation of milligram amounts of 21 deoxyribonucleic acid restriction fragments. Biochemistry. 1981 Jun 23;20(13):3748–3756. doi: 10.1021/bi00516a013. [DOI] [PubMed] [Google Scholar]
- Jack W. E., Terry B. J., Modrich P. Involvement of outside DNA sequences in the major kinetic path by which EcoRI endonuclease locates and leaves its recognition sequence. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4010–4014. doi: 10.1073/pnas.79.13.4010. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Langowski J., Pingoud A., Goppelt M., Maass G. Inhibition of Eco RI action by polynucleotides. A characterization of the non-specific binding of the enzyme to DNA. Nucleic Acids Res. 1980 Oct 24;8(20):4727–4736. doi: 10.1093/nar/8.20.4727. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Langowski J., Urbanke C., Pingoud A., Maass G. Transient cleavage kinetics of the Eco RI restriction endonuclease measured in a pulsed quench-flow apparatus: enzyme concentration-dependent activity change. Nucleic Acids Res. 1981 Jul 24;9(14):3483–3490. doi: 10.1093/nar/9.14.3483. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Richter P. H., Eigen M. Diffusion controlled reaction rates in spheroidal geometry. Application to repressor--operator association and membrane bound enzymes. Biophys Chem. 1974 Oct;2(3):255–263. doi: 10.1016/0301-4622(74)80050-5. [DOI] [PubMed] [Google Scholar]
- Riggs A. D., Bourgeois S., Cohn M. The lac repressor-operator interaction. 3. Kinetic studies. J Mol Biol. 1970 Nov 14;53(3):401–417. doi: 10.1016/0022-2836(70)90074-4. [DOI] [PubMed] [Google Scholar]
- Rubin R. A., Modrich P. Substrate dependence of the mechanism of EcoRI endonuclease. Nucleic Acids Res. 1978 Aug;5(8):2991–2997. doi: 10.1093/nar/5.8.2991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schranner R., Richter P. H. Rate enhancement by guided diffusion. Chain length dependence of repressor-operator association rates. Biophys Chem. 1978 May;8(2):135–150. doi: 10.1016/0301-4622(78)80005-2. [DOI] [PubMed] [Google Scholar]
- Winter R. B., Berg O. G., von Hippel P. H. Diffusion-driven mechanisms of protein translocation on nucleic acids. 3. The Escherichia coli lac repressor--operator interaction: kinetic measurements and conclusions. Biochemistry. 1981 Nov 24;20(24):6961–6977. doi: 10.1021/bi00527a030. [DOI] [PubMed] [Google Scholar]
- Winter R. B., von Hippel P. H. Diffusion-driven mechanisms of protein translocation on nucleic acids. 2. The Escherichia coli repressor--operator interaction: equilibrium measurements. Biochemistry. 1981 Nov 24;20(24):6948–6960. doi: 10.1021/bi00527a029. [DOI] [PubMed] [Google Scholar]
- Woodhead J. L., Malcolm A. D. Non-specific binding of restriction endonuclease EcoR1 to DNA. Nucleic Acids Res. 1980 Jan 25;8(2):389–402. doi: 10.1093/nar/8.2.389. [DOI] [PMC free article] [PubMed] [Google Scholar]
