Carnitine palmitoyltransferase 2: New insights on the substrate specificity and implications for acylcarnitine profiling

BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR BASIS OF DISEASE

卷:1802

Article

Violante, Sara^1,2  Ijlst, Lodewijk²  van Lenthe, Henk²  de Almeida, Isabel Tavares¹  Wanders, Ronald J.²  Ventura, Fatima V.¹ 

[1] Univ Lisbon, Fac Farm, Metab & Genet Grp, Res Inst Med & Pharmaceut Sci,iMed UL, P-1649003 Lisbon, Portugal

[2] Univ Amsterdam, Acad Med Ctr, Dept Clin Chem & Pediat, Lab Genet Metab Dis, NL-1105 AZ Amsterdam, Netherlands

关键词: Acylcarnitine;    Carnitine palmitoyltransferase 2 (CPT2);    Long-chain acyl-CoA;    Fatty acid oxidation (FAO);    Mitochondrial Trifunctional Protein (MTP);   

DOI  :  10.1016/j.bbadis.2010.06.002

来源: Elsevier

【 摘 要 】

Over the last years acylcarnitines have emerged as important biomarkers for the diagnosis of mitochondrial fatty acid beta-oxidation (mFAO) and branched-chain amino acid oxidation disorders assuming they reflect the potentially toxic acyl-CoA species, accumulating intramitochondrially upstream of the enzyme block. However, the origin of these intermediates still remains poorly understood. A possibility exists that carnitine palmitoyltransferase 2 (CPT2), member of the carnitine shuttle, is involved in the intramitochondrial synthesis of acylcarnitines from accumulated acyl-CoA metabolites. To address this issue, the substrate specificity profile of CPT2 was herein investigated. Saccharomyces cerevisiae homogenates expressing human CPT2 were incubated with saturated and unsaturated C2-C26 acyl-CoAs and branched-chain amino acid oxidation intermediates. The produced acylcarnitines were quantified by ESI-MS/MS. We show that CPT2 is active with medium (C8-C12) and long-chain (C14-C18) acyl-CoA esters, whereas virtually no activity was found with short- and very long-chain acyl-CoAs or with branched-chain amino acid oxidation intermediates. Trans-2-enoyl-CoA intermediates were also found to be poor substrates for CPT2. Inhibition studies performed revealed that trans-2-C16:1-CoA may act as a competitive inhibitor of CPT2 (K-i of 18.8 mu M). The results obtained clearly demonstrate that CPT2 is able to reverse its physiological mechanism for medium and long-chain acyl-CoAs contributing to the abnormal acylcarnitines profiles characteristic of most mFAO disorders. The finding that trans-2-enoyl-CoAs are poorly handled by CPT2 may explain the absence of trans-2-enoyl-carnitines in the profiles of mitochondrial trifunctional protein deficient patients, the only defect where they accumulate, and the discrepancy between the clinical features of this and other long-chain mFAO disorders such as very long-chain acyl-CoA dehydrogenase deficiency. (C) 2010 Elsevier B.V. All rights reserved.

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