Alternations of Some Enzymological Characterizations of Site-mutagenized DNA Polymerase ?
- VernacularTitle:定位诱变产生的DNA聚合酶?某些特性的改变
- Author:
Nan LIU
- Publication Type:Journal Article
- Keywords:
DNA polymerase ? site-mutagenesis;
enzyme active site
- From:
Academic Journal of Second Military Medical University
1985;0(05):-
- CountryChina
- Language:Chinese
-
Abstract:
The enzymological characterizations of site-mutagenized rat recombinant DNA polymerase?, RQ182 and RQ183 were studied The phosphocellulose column chromatographies showed that the mutant and the wild DNA polymerases were all eluted by about 0.5 mol/ L KCI, but the denatured DNA-cellulose chromatographies showed that although the wild enzyme was eluted by 0.35 mol/L KCI, RQ182 and RQ183 were eluted by 0.55 and 0.45 mol/L KCI, respectively, indicating that the binding abilities to DNA of the mutant enzymes were increased. Km values for the substrate (dTTP)of the wild enzyme, RQ182 and RQ183 were determined as 38.5, 34.5 and 111.1 ?mol/L, respectively,and the Km values for the primer (oligo(dT)) were 1.28, 1.96 and 6.58 ?g/ml, respectively. The results showed that the affinities of RQ183 to the substrate and the primer were decreased dramatically. It is suggested that Arg182 and Arg183 were involved in the active site function of DNA polymerase ?, in binding to DNA template, in recognizing of primer, and in binding to and catalyzing of substrates of the enzyme.
