REGULATION OF FOLATE AND ONE-CARBON METABOLISM IN MAMMALIAN-CELLS .1.FOLATE METABOLISM IN CHINESE-HAMSTER OVARY CELLS EXPRESSING ESCHERICHIA-COLI OR HUMAN FOLYLPOLY-GAMMA-GLUTAMATE SYNTHETASE-ACTIVITY
Cb. Osborne et al., REGULATION OF FOLATE AND ONE-CARBON METABOLISM IN MAMMALIAN-CELLS .1.FOLATE METABOLISM IN CHINESE-HAMSTER OVARY CELLS EXPRESSING ESCHERICHIA-COLI OR HUMAN FOLYLPOLY-GAMMA-GLUTAMATE SYNTHETASE-ACTIVITY, The Journal of biological chemistry, 268(29), 1993, pp. 21657-21664
Chinese hamster ovary (CHO) cell transfectants expressing various leve
ls of human and Escherichia coli folylpoly-gamma-glutamate synthetase
(FPGS) activity and possessing different folylpolyglutamate chain leng
th distributions have been developed as models for folate and antifola
te metabolism. The synthesis of pteroyltriglutamate was sufficient for
normal cellular retention of folate and also overcame the phenotypic
requirement for purines and thymidine of AUXB1, a CHO cell mutant lack
ing FPGS activity and lacking folylpolyglutamates. Only low levels of
FPGS are required to enable cellular metabolism of folates to forms th
at are retained by mammalian cells. The higher levels found in mammali
an cells are required for the synthesis of the long chain polyglutamat
e derivatives characteristic of mammalian cells. At low medium folate
concentrations, folate accumulation by transfectants expressing human
FPGS was not responsive to FPGS levels as the limiting step in metabol
ism was beyond the triglutamate, the chain length required for retenti
on. The rate-limiting step in folate metabolism in cells expressing th
e E. coli enzyme was the conversion of diglutamate to triglutamate, an
d, at low FPGS levels, the E. coli enzyme was about 50-fold less effec
tive than the human FPGS in enabling cellular folate accumulation. The
se data suggest that cellular accumulation of any folate analog whose
mono- or diglutamate derivative is a poor substrate for FPGS would be
very responsive to the level of FPGS activity.