The fatty acid transport protein (FATP1) is a very long chain acyl-CoA synthetase
The fatty acid transport protein (FATP1) is a very long chain acyl-CoA synthetase
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DOI：
10.1074/jbc.274.51.36300
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发表时间：
1999-12-17
影响因子：
4.8
通讯作者：
Bernlohr, DA
中科院分区：
文献类型：
--
作者：
Coe, NR;Smith, AJ;Bernlohr, DA
小鼠脂肪酸转运蛋白（FATP1）的一级序列与超长链（C20-C26）酰基辅酶a合成酶的多基因家族非常相似，为了确定FATP1是否是长链酰基辅酶a合成酶，我们在COS1细胞中表达了FATP1- myc /His融合蛋白，并对其酶活性进行了分析。此外，在酰基辅酶a合成酶中保守的两个结构域发生了突变：在假定的活性位点（氨基酸249-254）上发生了6个氨基酸的替换，产生了突变体M1；在保守的c端结构域（氨基酸464-523）上发生了59个氨基酸的缺失，产生了突变体M2。免疫定位发现，FATP1-Myc/His表达形式分布在COS1细胞膜和胞内膜之间。表达野生型FATP1-Myc/His的COS1细胞显示木质素酰辅酶a合成酶活性（C24:0）与棕榈酰辅酶a合成酶活性（C16:0）的比值增加了8倍，这是非常长链酰基辅酶a合成酶的特征，而突变体M1和M2都没有催化活性。用镍基亲和层析法部分纯化了洗涤剂溶解的FATP1-Myc/His，发现其超长链酰基辅酶a特异性活性增加了10倍（C24:0/C16:0）。这些结果表明，FATP1是一种非常长的链酰基辅酶a合成酶，并表明促进哺乳动物脂肪酸摄取的潜在机制是通过酯化偶联内流。
The primary sequence of the murine fatty acid transport protein (FATP1) is very similar to the multigene family of very long chain (C20-C26) acyl-CoA synthetases, To determine if FATP1 is a long chain acyl coenzyme A synthetase, FATP1-Myc/His fusion protein was expressed in COS1 cells, and its enzymatic activity was analyzed. In addition, mutations were generated in two domains conserved in acyl-CoA synthetases: a 6-amino acid substitution into the putative active site (amino acids 249-254) generating mutant M1 and a 59-amino acid deletion into a conserved C-terminal domain (amino acids 464-523) generating mutant M2. Immunolocalization revealed that the FATP1-Myc/His forms were distributed between the COS1 cell plasma membrane and intracellular membranes. COS1 cells expressing wild type FATP1-Myc/His exhibited a 8-fold increase in the ratio of lignoceroyl-CoA synthetase activity (C24:0) to palmitoyl-CoA synthetase activity (C16:0), characteristic of very long chain acyl-CoA synthetases, whereas both mutant M1 and M2 were catalytically inactive. Detergent-solubilized FATP1-Myc/His was partially purified using nickel-based affinity chromatography and demonstrated a 10-fold increase in very long chain acyl-CoA specific activity (C24:0/C16:0). These results indicate that FATP1 is a very long chain acyl-CoA synthetase and suggest that a potential mechanism for facilitating mammalian fatty acid uptake is via esterification coupled influx.