Residues 45 and 404 in the murine reduced folate carrier may interact to alter carrier binding and mobility.
Residues 45 and 404 in the murine reduced folate carrier may interact to alter carrier binding and mobility.
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鼠还原叶酸载体中的残基 45 和 404 可能相互作用，改变载体结合和迁移率。
DOI：
10.1016/s0005-2736(03)00136-6
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
Goldman,IDavid
中科院分区：
文献类型：
--
作者：
Zhao,Rongbao;Wang,Yanhua;Gao,Feng;Goldman,IDavid
还原型叶酸载体（RFC）是一种易化转运蛋白，在还原型叶酸和抗叶酸剂进入细胞的过程中起着重要作用。以前的研究表明，突变的E45 K在第一个跨膜结构域（TMD），和K404 L在第11个TMD，产生选择性和相反的改变，在结合天然叶酸底物的鼠RFC。前一种突变通常与抗叶酸剂耐药性有关。本研究旨在通过比较稳定转染K404 E、E45 K或E45 K/K404 E载体的RFC无效细胞系的转运特性来确定这些位点之间是否可能存在相互作用。结果表明：（1）所有突变载体均插入细胞质膜。(2)在K404 E突变体中，与L1210细胞相比，5-甲酰四氢叶酸和5-甲基四氢叶酸的流入Kt显著增加，叶酸的流入Kt在很小程度上增加。然而，随着第二个E45 K突变的引入，这些叶酸的流入Kt恢复到E45 K细胞的那些，其保留了5-甲基四氢叶酸的野生型结合，并增强了5-甲酰四氢叶酸和叶酸的结合。(3)E45 K突变体的内流Vmax明显降低。引入第二个K404 E突变使该参数加倍，并且5-甲酰四氢叶酸的Vmax与Kt的比率恢复至野生型携带者的约50%，这与功能的实质性增加一致。(4)氯抑制野生型RFC，但E45 K突变体需要氯的活动。K404 E突变体也被氯抑制，但K404 E突变的引入降低了E45 K的氯依赖性。结果表明，有E45和K404残基之间的相互作用，在第一和第11 TMD，分别，但E45残基似乎是更占主导地位的结合和阴离子敏感性的决定因素。
The reduced folate carrier (RFC), a facilitative transporter, plays a major role in the delivery of reduced folates and antifolates into cells. Previous studies indicated that mutations of E45K in the first transmembrane domain (TMD), and K404L in the 11th TMD, produce selective and opposite alterations in binding of natural folate substrates to murine RFC. The former mutation is frequently associated with antifolate resistance. The current study was designed to determine whether there might be an interaction between these sites by comparing the transport properties of RFC-null cell lines stably transfected with K404E, E45K, or E45K/K404E carriers. These studies demonstrated that: (1) All mutant carriers were inserted into the plasma membrane. (2) In the K404E mutant, the influx Kt's for 5-formyltetrahydrofolate and 5-methyltetrahydrofolate were markedly increased, and to a much smaller extent folic acid, as compared to L1210 cells. However, with introduction of a second E45K mutation the influx Ktfor these folates reverted to those of the E45K cells which retained wild-type binding for 5-methyltetrahydrofolate and enhanced binding of 5-formyltetrahydrofolate and folic acid. (3) The influx Vmaxof the E45K mutant was markedly reduced. Introduction of the second K404E mutation doubled this parameter and the ratio of Vmaxto Ktfor 5-formytetrahydrofolate was restored to ∼50% that of the wild-type carrier consistent with a substantial increase in function. (4) Chloride inhibits wild-type RFC but the E45K mutant requires chloride for activity. The K404E mutant is also suppressed by chloride but introduction of the K404E mutation decreased the chloride-dependence of E45K. The results suggest that there is an interaction between the E45 and K404 residues in the first and 11th TMDs, respectively, but that the E45 residue appears to be the more dominant determinant of binding and anion sensitivity.
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N-糖基化能力丧失对人还原叶酸载体的转运活性和细胞定位的影响。
DOI：
--
发表时间：
1998
期刊：
Biochimica et Biophysica Acta
影响因子：
--
作者：
S. Wong;L. Zhang;S. A. Proefke;L. Matherly
通讯作者：
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氨基酸 45 残基在降低叶酸载体功能和离子依赖性转运中的作用，以定点诱变为特征。
影响因子：
3.6
作者：
Zhao,R;Gao,F;Wang,PJ;Goldman,ID
通讯作者：
Goldman,ID
通过高压液相色谱在体外艾氏腹水肿瘤细胞中评估甲氨蝶呤聚-γ-谷氨酰衍生物的快速形成及其与二氢叶酸还原酶的关联。
DOI：
--
发表时间：
1982
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Fry,DW;Yalowich,JC;Goldman,ID
通讯作者：
Goldman,ID
L1210 鼠白血病细胞中的还原叶酸载体是一种 58 kDa 的蛋白质。
DOI：
10.1016/s0005-2736(00)00190-5
发表时间：
2000
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
Zhao,R;Gao,F;Liu,L;Goldman,ID
通讯作者：
Goldman,ID
影响因子：
4.8
作者：
Jansen, G;Mauritz, R;Assaraf, YG
通讯作者：
Assaraf, YG