The restriction endonuclease BstI was purified from 70kg of Bacillus stearothermophilus. The final product is at least 97% pure as judged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis; this major protein species co-migrates with the enzyme activity on native polyacrylamide-gel electrophoresis and isoelectric focusing. Pure restriction endonuclease BstI has a subunit mol.wt. of 26,000 and is probably a loosely associated dimer. The enzyme shows maximum activity at pH values between 7 and 9.5, and in the presence of 0.5-2mM-Mg2+. NaCl inhibits the restriction enzyme activity. Restriction endonuclease BstI cleaves DNA in a position identical with that cleaved by endonuclease BamHI (for Bacillus amyloliquefaciens), i.e.: (formula: see text). In the presence of high concentrations of enzyme, DNA cleavage occurs at secondary sites. This side-specificity is enhanced by the addition of glycerol. Preliminary studies indicate that these sites are of the type: (formula: see text).
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January 1979
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Research Article|
January 01 1979
Purification, properties and specificity of the restriction endonuclease from Bacillus stearothermophilus
Publisher: Portland Press Ltd
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 1979 London: The Biochemical Society
1979
Biochem J (1979) 177 (1): 49–62.
Citation
C M Clarke, B S Hartley; Purification, properties and specificity of the restriction endonuclease from Bacillus stearothermophilus. Biochem J 1 January 1979; 177 (1): 49–62. doi: https://doi.org/10.1042/bj1770049
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