The mitochondrial carnitine/acylcarnitine carrier (CAC) is characterized by the presence of a distinct motif, RXXPANAAXF, within its sixth transmembrane alpha-helix. In this study, we analysed the role of the amino acids of this motif in the structure-function relationships of the human CAC by using two complementary approaches. First, we performed functional analysis in the model fungus Aspergillus nidulans of selected mutations with structural and functional relevance. Second, similar mutant human CACs were biochemically characterized after their reconstitution into liposomes. Both analyses have provided relevant information on the importance and role of the CAC motif residues in the activity and metabolic function of CAC. Only the two adjacent alanines, Ala281 and Ala282 in the human CAC, have been found not to be crucial for transport activity and in vivo function. Results obtained from amino acid substitutions of residues Arg275, Asn280 and Phe284 of human CAC together with structural analysis using molecular modelling of the carrier suggest that R275, N280 and F284 are involved in substrate binding during acylcarnitine/carnitine translocation. Furthermore, functional analysis of mutations of residues Pro278 and Ala279 in A. nidulans, together with kinetic data in reconstituted liposomes, suggest a predominant structural role for these amino acids.

中文翻译：

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肉碱/酰基肉碱转位酶RX2PANAAXF独特基序内的残基功能表征。

线粒体肉碱/酰基肉碱载体（CAC）的特征是在其第六个跨膜α-螺旋中存在独特的基序RXXPANAAXF。在这项研究中，我们通过使用两种互补方法分析了该基序氨基酸在人类CAC的结构-功能关系中的作用。首先，我们在具有结构和功能相关性的选定突变的模型真菌构巢曲霉中进行了功能分析。第二，相似的突变型人CACs重组为脂质体后，经过生物化学鉴定。两种分析都提供了有关CAC模序残基在CAC活性和代谢功能中的重要性和作用的相关信息。人类CAC中只有两个相邻的丙氨酸Ala281和Ala282，已经发现对于转运活性和体内功能不是至关重要的。从人CAC残基Arg275，Asn280和Phe284的氨基酸置换以及使用载体分子模型进行的结构分析获得的结果表明，R275，N280和F284在酰基肉碱/肉碱易位期间参与底物结合。此外，构巢曲霉（A.nidulans）中残基Pro278和Ala279的突变的功能分析以及重组脂质体中的动力学数据表明这些氨基酸的主要结构作用。N280和F284在酰基肉碱/肉碱易位期间参与底物结合。此外，构巢曲霉（A.nidulans）中残基Pro278和Ala279的突变的功能分析以及重组脂质体中的动力学数据表明这些氨基酸的主要结构作用。N280和F284在酰基肉碱/肉碱易位期间参与底物结合。此外，构巢曲霉（A.nidulans）中残基Pro278和Ala279的突变的功能分析以及重组脂质体中的动力学数据表明这些氨基酸的主要结构作用。