Identification of conserved amino acid residues in rat liver carnitine palmitoyltransferase I critical for malonyl-CoA inhibition. Mutation of methionine 593 abolishes malonyl-CoA inhibition
Casals i Farré, Núria
Hegardt, Fausto G.
Carnitine palmitoyltransferase (CPT) I, which catalyzes the conversion of palmitoyl-CoA to palmitoylcarnitine facilitating its transport through the mitochondrial membranes, is inhibited by malonyl-CoA. By using the SequenceSpace algorithm program to identify amino acids that participate in malonyl-CoA inhibition in all carnitine acyltransferases, we found 5 conserved amino acids (Thr314, Asn464, Ala478, Met593, and Cys608, rat liver CPT I coordinates) common to inhibitable malonyl-CoA acyltransferases (carnitine octanoyltransferase and CPT I), and absent in noninhibitable malonyl-CoA acyltransferases (CPT II, carnitine acetyltransferase (CAT) and choline acetyltransferase (ChAT)). To determine the role of these amino acid residues in malonyl-CoA inhibition, we prepared the quintuple mutant CPT I T314S/N464D/A478G/M593S/C608A as well as five single mutants CPT I T314S, N464D, A478G, M593S, and C608A. In each case the CPT I amino acid selected was mutated to that present in the same homologous position in CPT II, CAT, and ChAT. Because mutant M593S nearly abolished the sensitivity to malonyl-CoA, two other Met593mutants were prepared: M593A and M593E. The catalytic efficiency (V max/K m) of CPT I in mutants A478G and C608A and all Met593 mutants toward carnitine as substrate was clearly increased. In those CPT I proteins in which Met593 had been mutated, the malonyl-CoA sensitivity was nearly abolished. Mutations in Ala478, Cys608, and Thr314 to their homologous amino acid residues in CPT II, CAT, and ChAT caused various decreases in malonyl-CoA sensitivity. Ala478 is located in the structural model of CPT I near the catalytic site and participates in the binding of malonyl-CoA in the low affinity site (Morillas, M., Gómez-Puertas, P., Rubı́, B., Clotet, J., Ariño, J., Valencia, A., Hegardt, F. G., Serra, D., and Asins, G. (2002) J. Biol. Chem. 277, 11473–11480). Met593 may participate in the interaction of malonyl-CoA in the second affinity site, whose location has not been reported.
57 - Biologia
61 - Medicina
Metabolisme -- Trastorns
Aminoàcids -- Metabolisme
Metabolismo -- Trastornos
Aminoácidos -- Metabolismo
Metabolism -- Disorders
Amino acids -- Metabolism
American Society for Biochemistry and Molecular Biology
Is part of
Journal of Biological Chemistry
Morillas, Montserrat; Gómez-Puertas, Paulino; Bentebibel, Assia [et al.]. Identification of conserved amino acid residues in rat liver carnitine palmitoyltransferase I critical for malonyl-CoA inhibition. Mutation of methionine 593 abolishes malonyl-CoA inhibition. Journal of Biological Chemistry, 2003, 278(11), p. 9058-9063. Disponible en: <https://www.sciencedirect.com/science/article/pii/S0021925819712745?via%3Dihub>. Fecha de acceso: 5 may. 2021. DOI: 10.1074/jbc.M209999200
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