title: The type IIs restriction endonuclease BspMI is a tetramer that acts concertedly at two copies of an asymmetric DNA sequence.
abstract: Type IIs endonucleases recognize asymmetric DNA sequences and cleave both strands at fixed positions downstream of the sequence. Many type IIs enzymes, including BspMI, cleave substrates with two sites more rapidly than those with one site. They usually act sequentially on DNA with two sites, but BspMI converted such a substrate directly to the final products cut at both sites. The BspMI endonuclease was found to be a tetramer, in contrast to the monomeric structures for many type IIs enzymes. No change in subunit association occurred during the BspMI reaction. Plasmids with two BspMI sites were cleaved in cis, in reactions spanning sites in the same DNA, even when the sites were separated by just 38 bp. Plasmids with one BspMI site were cleaved in trans, with the enzyme bridging sites in separate DNA molecules: these slow reactions could be accelerated by adding a second DNA with the recognition sequence. Thus, whereas many type IIs enzymes dimerize before cleaving DNA, a process facilitated by two recognition sites in cis, the BspMI tetramer binds two copies of its recognition sequence before cleaving the DNA in both strands at both sites.
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, University Walk, Bristol BS8 1TD, United Kingdom.
authors.forename: Niall A
authors.initials: NA
authors.lastname: Gormley
authors.forename: Anna L
authors.initials: AL
authors.lastname: Hillberg
authors.forename: Stephen E
authors.initials: SE
authors.lastname: Halford
mesh_terms: D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D007700:Kinetics; D012492:Salts; D013379:Substrate Specificity; D014461:Ultracentrifugation
chemical_list: D001704:Biopolymers; D012492:Salts; D004247:DNA; C052649:endodeoxyribonuclease BspMI; D015252:Deoxyribonucleases, Type II Site-Specific