Functional characterization of PCFT/HCP1 as the molecular entity of the carrier-mediated intestinal folate transport system in the rat model
Functional characterization of PCFT/HCP1 as the molecular entity of the carrier-mediated intestinal folate transport system in the rat model
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DOI：
10.1152/ajpgi.00309.2007
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发表时间：
2008-03-01
影响因子：
4.5
通讯作者：
Yuasa, Hiroaki
中科院分区：
文献类型：
--
作者：
Inoue, Katsuhisa;Nakai, Yasuhiro;Yuasa, Hiroaki
质子偶联叶酸转运蛋白/血红素载体蛋白1（PCFT/HCP 1）是近年来发现的一种通过质子偶联机制介导叶酸跨膜转运的转运蛋白，可能是载体介导的肠道叶酸转运系统的分子实体。为了进一步阐明其在肠道叶酸转运中的作用，我们研究了在非洲爪蟾卵母细胞中表达的大鼠PCFT/HCP 1（rPCFT/HCP 1）的功能特性，并与使用外翻组织囊评价的大鼠小肠中载体介导的叶酸转运系统的功能特性进行了比较。证明rPCFT/HCP 1在较低的酸性pH下更有效地转运叶酸和甲氨蝶呤，并且在pH 5.5下评估，前者的米氏常数（Km）（2.4 μ M）小于后者（5.7 μ M），表明其作为质子偶联叶酸转运蛋白的特征，其有利于叶酸而不是甲氨蝶呤作为底物。发现rPCFT/HCP 1介导的叶酸转运被几种但有限的阴离子化合物抑制，如磺溴酞和柳氮磺胺吡啶。rPCFT/HCP 1的这些特性与叶酸载体介导的小肠转运系统一致，其在大鼠小肠中对叶酸和甲氨蝶呤的Km值分别为1.2和5.8 μ M。叶酸转运系统活性沿着肠道的分布与rPCFT/HCP 1 mRNA的分布一致。这项研究是第一个克隆rPCFT/HCP 1，我们成功地提供了几条证据，表明其作为肠道叶酸转运系统的分子实体的作用。
Proton-coupled folate transporter/heme carrier protein 1 (PCFT/HCP1) has recently been identified as a transporter that mediates the translocation of folates across the cellular membrane by a proton-coupled mechanism and suggested to be the possible molecular entity of the carrier-mediated intestinal folate transport system. To further clarify its role in intestinal folate transport, we examined the functional characteristics of rat PCFT/HCP1 (rPCFT/HCP1) expressed in Xenopus laevis oocytes and compared with those of the carrier-mediated folate transport system in the rat small intestine evaluated by using the everted tissue sacs. rPCFT/HCP1 was demonstrated to transport folate and methotrexate more efficiently at lower acidic pH and, as evaluated at pH 5.5, with smaller Michaelis constant (Km) for the former (2.4 mu M) than for the latter (5.7 mu M), indicating its characteristic as a proton-coupled folate transporter that favors folate than methotrexate as substrate. rPCFT/HCP1-mediated folate transport was found to be inhibited by several but limited anionic compounds, such as sulfobromophthalein and sulfasalazine. All these characteristics of rPCFT/HCP1 were in agreement with those of carrier-mediated intestinal folate transport system, of which the Km values were 1.2 and 5.8 mu M for folate and methotrexate, respectively, in the rat small intestine. Furthermore, the distribution profile of the folate transport system activity along the intestinal tract was in agreement with that of rPCFT/HCP1 mRNA. This study is the first to clone rPCFT/HCP1, and we successfully provided several lines of evidence that indicate its role as the molecular entity of the intestinal folate transport system.