4EPH: CRYSTAL STRUCTURE OF RAT CARNITINE PALMITOYLTRANSFERASE 2 IN COMPLEX with CoA-site inhibitor

Citation:
Abstract
Carnitine palmitoyl transferase 2 (CPT-2) is a key enzyme in the mitochondrial fatty acid metabolism. The active site is comprised of a Y-shaped tunnel with distinct binding sites for the substrate acylcarnitine and the cofactor CoA. We investigated the thermodynamics of binding of four inhibitors directed against either the CoA or the acylcarnitine binding sites using isothermal titration calorimetry (ITC). CPT-2 is a monotopic membrane protein and was solubilized by beta-octylglucoside (beta-OG) above its critical micellar concentration (CMC) to perform inhibitor titrations in solutions containing detergent micelles. The CMC of beta-OG in the presence of inhibitors was measured with ITC and small variations were observed. The inhibitors bound to rat CPT-2 (rCPT-2) with 1:1 stoichiometry and the dissociation constants were in the range of K D = 2-20 muM. New X-ray structures and docking models of rCPT-2 in complex with inhibitors enable an analysis of the thermodynamic data in the context of the interaction observed for the individual binding sites of the ligands. For all ligands the binding enthalpy was exothermic, and enthalpy as well as entropy contributed to the binding reaction, with the exception of ST1326 for which binding was solely enthalpy-driven. The substrate analog ST1326 binds to the acylcarnitine binding site and a heat capacity change close to zero suggests a balance of electrostatic and hydrophobic interactions. An excellent correlation of the thermodynamic (ITC) and structural (X-ray crystallography, models) data was observed suggesting that ITC measurements provide valuable information for optimizing inhibitor binding in drug discovery.
PDB ID: 4EPHDownload
MMDB ID: 109175
PDB Deposition Date: 2012/4/17
Updated in MMDB: 2017/11
Experimental Method:
x-ray diffraction
Resolution: 2.3  Å
Source Organism:
Similar Structures:
Biological Unit for 4EPH: monomeric; determined by author and by software (PISA)
Molecular Components in 4EPH
Label Count Molecule
Protein (1 molecule)
1
Carnitine O-palmitoyltransferase 2, Mitochondrial(Gene symbol: Cpt2)
Molecule annotation
Chemicals (2 molecules)
1
1
2
1
* Click molecule labels to explore molecular sequence information.

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