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PMID: 3889850 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

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

Nucleic acids research ·Vol. 13 ·No. 8 ·1985-04-25 ·Pages 2921-30

Chu G, Berg P

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.

MeSH Terms
Animals Chromatography, Thin Layer Escherichia coli/enzymology,genetics Methods Mice Pentosyltransferases/analysis Ribonucleosides/metabolism Ribonucleotides/metabolism Transduction, Genetic Xanthine Xanthines/metabolism
Chemicals
Ribonucleosides Ribonucleotides Xanthines Xanthine xanthosine monophosphate xanthosine Pentosyltransferases xanthine phosphoribosyltransferase
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Chu G
Berg P
References (16)
16 references, click to expand
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/13.8.2921
Published
1985-04-25
Pages
2921-30
Language
English
Region
England
NLM ID
0411011
PMCID
PMC341204
Subset
IM
Grants
NIGMS NIH HHS · GM-13235 · United States
External Links
PubMed source DOI: 10.1093/nar/13.8.2921 PMC Full Text View journal details
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