Age-dependent expression of a novel protein in mouse liver immunologically and functionally homologous with dihydrofolate reductase.
Dihydrofolate reductase with a molecular weight of 22,000 has been purified by salt precipitation and methotrexate-Sepharose affinity chromatography from mouse livers having a mean weight of 2.4 g each. When the same purification procedure was followed using livers with a mean weight of 1.4 g or less, a protein with a molecular weight of 27,500 co-purified with dihydrofolate reductase. This 27.5 kDa species was recovered with dihydrofolate reductase following a second passage through the affinity column and it reacted by Western immunoblotting with a monospecific polyclonal antiserum raised to the purified 22 kDa enzyme. The two proteins could not be separated in a native state to compare their functional activity, but the 27.5 kDa protein appeared to have catalytic reductase activity when assayed directly on the polyacrylamide gel following non-denaturing electrophoresis. The catalytic activity of the mixture of the purified proteins was stoichiometrically inhibited by a molar equivalent of methotrexate.
Kyner
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Department of Medicine, Veterans Hospital, Brooklyn, NY.
Rothenberg
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SP
D000375:Aging; D000818:Animals; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology / Q000254:growth & development*; D008727:Methotrexate / Q000378:metabolism / Q000494:pharmacology; D051379:Mice; D008815:Mice, Inbred Strains; D008970:Molecular Weight; D013762:Tetrahydrofolate Dehydrogenase / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism
D007527:Isoenzymes; D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate