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Binding of non-substrate nucleotides to a restriction endonuclease: a model for the interaction of bam HI with its recognition sequence.
Binding of non-substrate nucleotides to a restriction endonuclease: a model for the interaction of bam HI with its recognition sequence.
- Source :
-
Nucleic acids research [Nucleic Acids Res] 1980 Jun 11; Vol. 8 (11), pp. 2547-59. - Publication Year :
- 1980
-
Abstract
- The kinetic constants of the site-specific endonuclease Bam HI for various substrates were determined and binding of non-substrate nucleotides to the enzyme was studied. Agarose gel assays in combination with an integrated Michaelis-Menten equation were used for the evaluation of data. The turnover number was 2.2 min-1 at 37 degrees C with pJC80 DNA as the substrate. It depends on the conformation and base composition of the substrate. Michaelis constants also depend on substrate conformation. Non-substrate polynucleotides were found to inhibit Bam competitively with KI ranging from 10(-6) to > 10(-3) M depending on base composition, base pairing, and helix conformation. Dinucleotides showed sequence-specific, competitive inhibition with KIs ranging from 10(-5) to > 10(-3) M. Mononucleotides and -nucleosides acted noncompetitively. Binding was influenced by the extent of phosphorylation, but not by the nature of the base. KIs varied between 10(-3) and 10(-2) M. The results are discussed with respect to the recognition requirements of Bam HI.
- Subjects :
- Bacteriophage lambda metabolism
Bacteriophages metabolism
Base Composition
Base Sequence
DNA, Viral metabolism
Deoxyribonuclease BamHI
Escherichia coli metabolism
Kinetics
Nucleic Acid Conformation
RNA, Bacterial metabolism
RNA, Fungal metabolism
RNA, Viral metabolism
Saccharomyces cerevisiae metabolism
Structure-Activity Relationship
DNA Restriction Enzymes metabolism
Nucleotides metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0305-1048
- Volume :
- 8
- Issue :
- 11
- Database :
- MEDLINE
- Journal :
- Nucleic acids research
- Publication Type :
- Academic Journal
- Accession number :
- 6160464
- Full Text :
- https://doi.org/10.1093/nar/8.11.2547