Rapid assay for detection of Escherichia coli xanthine-guanine phosphoribosyltransferase activity in transduced cells.

Abstract

Cultured mammalian cells transduced with the Escherichia coli gene, Ecogpt, synthesize the bacterial enzyme xanthine-guanine phosphoribosyl transferase (XGPT) (1). This paper describes a method for measuring XGPT activity in crude cell extracts by following the conversion of 14C-xanthine (X) to 14C-xanthine monophosphate (XMP) and 14C-xanthosine (XR) by thin layer chromatography. The method is rapid, easy to use, sensitive and linear over a wide range of XGPT activity and has been useful for detecting XGPT in cells that were transiently transfected or stably transformed with Ecogpt. During our studies, we have found that a human cell line (XP20S) converts xanthine to XMP. This activity is probably catalyzed by a variant hypoxanthine-guanine phosphoribosyltransferase (HGPT) since the low activity is readily inhibited by hypoxanthine. A low level of conversion of X to XMP may explain why some cell lines are not killed in a medium containing mycophenolic acid and X.

Cite this paper

@article{Chu1985RapidAF, title={Rapid assay for detection of Escherichia coli xanthine-guanine phosphoribosyltransferase activity in transduced cells.}, author={Gilbert Chu and Paul Berg}, journal={Nucleic acids research}, year={1985}, volume={13 8}, pages={2921-30} }