Purification and characterization of Escherichia coli xanthine-guanine phosphoribosyltransferase produced by plasmid pSV2gpt.
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Deo SS, Tseng WC, Saini R, Coles RS, Athwal RS
Purification and characterization of Escherichia coli xanthine-guanine phosphoribosyltransferase produced by plasmid pSV2gpt.
Biochim Biophys Acta. 1985 May 8;839(3):233-9.
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- 3886014 [ View in PubMed]
- Abstract
The enzyme xanthine-guanine phosphoribosyltransferase from Escherichia coli cells harboring the plasmid pSV2gpt has been purified 30-fold to near homogeneity by single-step GMP-agarose affinity chromatography. It has a Km value of 2.5, 42 and 182 microM for the substrates guanine, xanthine and hypoxanthine, respectively, with guanine being the most preferred substrate. The enzyme exhibits a Km value of 38.5 microM for PRib-PP with guanine as second substrate and of 100 microM when xanthine is used as the second substrate. It is markedly inhibited by 6-thioguanine, GMP and to a lesser extent by some other purine analogues. Thioguanine has been found to be the most potent inhibitor. The subunit molecular weight of xanthine-guanine phosphoribosyltransferase was determined to be 19 000. The in situ activity assay on a nondenaturing polyacrylamide gel electrophoresis gel has indicated that a second E. coli phosphoribosyltransferase preferentially uses hypoxanthine as opposed to guanine as a substrate, and it does not use xanthine.