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A COMPARISON OF THYMIDYLATE SYNTHETASE ACTIVITIES FROM 5‐FLUORODEOXYURIDINE SENSITIVE AND RESISTANT VARIANTS OF MOUSE NEUROBLASTOMA 1
Author(s) -
Baskin F.,
Rosenberg R. N.
Publication year - 1975
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1975.tb06958.x
Subject(s) - thymidylate synthase , enzyme , chinese hamster , biochemistry , microbiology and biotechnology , biology , methyltransferase , cell culture , enzyme assay , substrate (aquarium) , methylation , in vitro , dna , fluorouracil , chemotherapy , ecology , genetics
— We have previously described a 5‐fluorodeoxyuridine (FUdR) resistant variant of mouse neuroblastoma possessing an 8‐fold elevation in methylenetetrahydrofolate: dUrd‐5′‐P C‐methyltransferase (EC2.1.1.b) [trivial name: thymidylate synthetase] specific activity relative to that of the sensitive parental clone. This increased specific activity is not associated with a change in cytoplasmic inhibitors or activators, a decreased degradation rate of the enzyme, or the synthesis of a new species with an increased V max , but appears to represent an increased synthesis of the enzyme species found in the sensitive parental clone. More resistant cell populations demonstrate even higher specific activities of this enzyme. The enzymatic activities from both the FUdR sensitive and resistant cells have identical stabilities to sonication, freezing, heat, urea, and sodium dodecyl sulfate, are equally and strongly inhibited by 5‐fluorodeoxyuridine‐5′‐phosphate, and have the same affinity for the substrate 2′‐deoxyuridine‐5′‐phosphate ( K m = 1·4 = 10 −6 m ). Both are stimulated by the addition of mercaptans and partially protected from heat denaturation in the presence of substrate. Unlike Don Chinese hamster cells (C onrad & R uddle 1972) an actinomycin d pulse of neuroblastoma cells in monolayer culture did not increase the thymidylate synthetase specific activity. Mixed growth of FUdR sensitive and resistant cells produced only additive activities.