Rapid assay for detection of Eschenchia coli xanthine-guanine phosphoribosyltransferase activity in transduced cells (original) (raw)

Journal Article

Gilbert Chu ,

Biochemistry Department, Stanford University Medical Center

Stanford, CA 94305, USA

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Paul Berg

Biochemistry Department, Stanford University Medical Center

Stanford, CA 94305, USA

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Received:

19 February 1985

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Gilbert Chu, Paul Berg, Rapid assay for detection of Eschenchia coli xanthine-guanine phosphoribosyltransferase activity in transduced cells, Nucleic Acids Research, Volume 13, Issue 8, 25 April 1985, Pages 2921–2930, https://doi.org/10.1093/nar/13.8.2921
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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 (XP2OS) 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.

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