Structural model of carnitine palmitoyltransferase I based on the carnitine acetyltransferase crystal

Abstract

CPT I (carnitine palmitoyltransferase I) catalyses the conversion of palmitoyl-CoA into palmitoylcarnitine in the presence of L-carnitine, facilitating the entry of fatty acids into mitochondria. We propose a 3-D (three-dimensional) structural model for L-CPT I (liver CPT I), based on the similarity of this enzyme to the recently crystallized mouse carnitine acetyltransferase. The model includes 607 of the 773 amino acids of L-CPT I, and the positions of carnitine, CoA and the palmitoyl group were assigned by superposition and docking analysis. Functional analysis of this 3-D model included the mutagenesis of several amino acids in order to identify putative catalytic residues. Mutants D477A, D567A and E590D showed reduced L-CPT I activity. In addition, individual mutation of amino acids forming the conserved Ser685-Thr686-Ser687 motif abolished enzyme activity in mutants T686A and S687A and altered K(m) and the catalytic efficiency for carnitine in mutant S685A. We conclude that the catalytic residues are His473 and Asp477, while Ser687 probably stabilizes the transition state. Several conserved lysines, i.e. Lys455, Lys505, Lys560 and Lys561, were also mutated. Only mutants K455A and K560A showed decreases in activity of 50%. The model rationalizes the finding of nine natural mutations in patients with hereditary L-CPT I deficiencies.

Carnitine palmitoyltransferase IA polymorphism P479L is common in Greenland Inuit and is associated with elevated plasma apolipoprotein A-I

“…These residues are in close proximity to Pro-479, and because the P479L mutant is the only natural mutant to diminish malonyl-CoA sensitivity while retaining catalytic function, the region from Ala-478 to His-483 has been proposed as a malonyl-CoA coupling domain (16). The model structure of CPT1A based on the carnitine acetyltransferase crystal places this domain close to carnitine and acyl-CoA binding sites as well as the proposed malonyl-CoA binding site (16,18,19).…”

C75 is converted to C75-CoA in the hypothalamus, where it inhibits carnitine palmitoyltransferase 1 and decreases food intake and body weight

“…CrAT D356A/M564G was modelled by the same procedures using rat CrAT wt as template. CPT1A was also modelled using as templates the structures of mouse carnitine octanoyltransferase (1XL7, 1XL8) [29] and carnitine palmitoyltransferase 2 (PDB entry 2H4T) [30], essentially as described elsewhere [31,32]. The quality of the models was checked using the WHAT-CHECK routines [33] from the WHAT-IF program [34] and the PROCHECK validation program from the SWISS-MODEL server facilities [35].…”

Carnitine palmitoyltransferase 1C: From cognition to cancer

“…No crystal structures are available for these two enzymes owing to the fact that they are integral membrane proteins and they lose catalytic function when solubilized. However, homology analysis of their sequences compared to those of the globular, soluble members of the carnitine acyltransferase family (CrAT, COT, CPT2) for which crystal structures have been obtained at a high resolution as well as homology modelling and docking using highly similar sequence motifs identified in other proteins have allowed in silico models of CPT1A tertiary structure [5][6][7]. Moreover, extensive structure-function relationship studies of CPT1A have provided evidence of intricate interactions between a relatively small (47 residues) regulatory N-terminal domain and a large (approx.…”