Mutagenesis of the DNA binding residues in bovine pancreatic DNase 1: an investigation into the mechanism of sequence discrimination by a sequence selective nuclease.
The sequence selectivity of DNase 1 cleavage has been investigated by site-directed mutagenesis of a chemically synthesised gene. Two key DNA binding residues have been conservatively altered (Y76F and R41K) or have had their side-chains truncated (Y76A and R41A) and the effect on the cleavage of tyr T promoter DNA has been noted. It would appear from these studies that DNase 1 is not sensitive to minor groove width via these DNA-contacting residues, and it is suggested that DNA helical stiffness is a controlling parameter in determining DNase 1 sequence selectivity.
Doherty
A J
AJ
Department of Biochemistry (SERC Molecular recognition Initiative Centre), University of Southampton, UK.
Worrall
A F
AF
Connolly
B A
BA
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics*; D003850:Deoxyribonuclease I / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D016296:Mutagenesis; D010179:Pancreas / Q000201:enzymology*; D011401:Promoter Regions, Genetic; D013379:Substrate Specificity
D004268:DNA-Binding Proteins; D004247:DNA; D003850:Deoxyribonuclease I