Deficient synthesis of MTHFD, a trifunctional folate-dependent enzyme, in the CHO Ade E mutant.
MTHFD is a folate-dependent trifunctional protein comprised of three activities: N5,N10-methylenetetrahydrofolate dehydrogenase, N5,N10-methenyltetrahydrofolate cyclohydrolase, and N10-formyltetrahydrofolate synthetase. The enzymes catalyze sequential interconversion of tetrahydrofolate derivatives required for purine, methionine, and thymidylate synthesis. A Chinese hamster ovary cell line (Ade-E), reported to have reduced cyclohydrolase activity, was studied to characterize the nature of the mutation. Enzymatic assays showed reduced activities of all three enzymes of the polypeptide. Immunoblotting and immunoprecipitation of radiolabeled cell extracts indicated that MTHFD protein was greatly reduced or absent in the mutant. Northern analysis of a clonal derivative of Ade-E revealed normal levels of MTHFD mRNA. These results suggest that the mutation affects a posttranscriptional process in the synthesis of the trifunctional enzyme.
Mascisch
A
A
Department of Pediatrics & Biology, McGill University-Montreal Children's Hospital Research Institute, Quebec.
Rozen
R
R
D000582:Amidophosphoribosyltransferase / Q000172:deficiency* / Q000235:genetics; D000619:Aminohydrolases / Q000096:biosynthesis / Q000172:deficiency* / Q000235:genetics; D000818:Animals; D015139:Blotting, Southern; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D005260:Female; D005492:Folic Acid / Q000502:physiology*; D005574:Formate-Tetrahydrofolate Ligase / Q000172:deficiency* / Q000235:genetics; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP) / Q000096:biosynthesis / Q000172:deficiency* / Q000235:genetics; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000172:deficiency* / Q000235:genetics; D009154:Mutation; D010053:Ovary / Q000201:enzymology*; D012323:RNA Processing, Post-Transcriptional
D009097:Multienzyme Complexes; C021836:formyl-methenyl-methylenetetrahydrofolate synthetase; D005492:Folic Acid; D008754:Methylenetetrahydrofolate Dehydrogenase (NADP); D000582:Amidophosphoribosyltransferase; D000619:Aminohydrolases; D005574:Formate-Tetrahydrofolate Ligase