Dipartimento di Genetica e Microbiologia "A. Buzzati Traverso", Sezione di Microbiologia e Fisiologia Vegetale, Università di Pavia, Via S. Epifanio 14, 27100, Pavia, Italy.
Plant Mol Biol. 1988 Jul;10(4):331-8. doi: 10.1007/BF00029883.
Dihydrofolate reductase (DHFR) and thymidylate synthase (TS) activities from cell suspension cultures of Daucus carota were shown to copurify on (NH4)2SO4 fractionation, DEAE Sephadex and methotrexate-Sepharose affinity chromatography and to share approximately the same Mr(183 kDa and 185 kDa respectively) as judged by gel filtration on Sephacryl S-200.The copurified protein migrated as a single band on polyacrylamide gel electrophoresis under denaturing conditions.Both activities could be eluted from the same position of the native gel.Moreover, methotrexate-resistant cell lines which overproduce DHFR revealed to have a parallel higher level of TS. It is therefore proposed and discussed that in carrot, similarly to protozoa, TS and DHFR are present on a single bifunctional polypeptide of 58 kDa.
细胞悬浮培养的胡萝卜二氢叶酸还原酶（DHFR）和胸苷酸合成酶（TS）的活性在（NH4）2SO4分级分离、DEAE Sephadex 和氨甲喋呤-Sepharose 亲和层析时被证明能共纯化，并且根据 Sephacryl S-200 的凝胶过滤，其分子量（Mr）分别约为 183 kDa 和 185 kDa。共纯化的蛋白质在变性条件下的聚丙烯酰胺凝胶电泳中迁移为单一条带。两种活性都可以从天然凝胶的相同位置洗脱。此外，对氨甲喋呤具有抗性并过量产生 DHFR 的细胞系显示出 TS 的水平也相应升高。因此，有人提出并讨论，在胡萝卜中，与原生动物类似，TS 和 DHFR 存在于一个 58 kDa 的单一双功能多肽上。
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