title: Oxidative inactivation of a charge transfer complex in the medium-chain acyl-CoA dehydrogenase.
abstract: The intense charge transfer complex between the enolate of 3-thia-octanoyl-CoA and the oxidized flavin of the medium-chain acyl-CoA dehydrogenase is discharged by the ferricenium ion with irreversible inactivation of the enzyme. Charge transfer complex formation is a necessary, but insufficient, condition for oxidative inactivation: the 3-oxa-octanoyl-CoA complex is also inactivated, whereas the comparable trans-3-octenoyl-CoA species is not. Complete inactivation of the dehydrogenase with 3-thia-octanoyl-CoA requires 1 molecule of thioester and apparently 3 molecules of ferricenium hexafluorophosphate. Experiments with 8-Cl-FAD substituted enzyme and the crystal structure of enzyme.ligand complexes argue that ferricenium ion-mediated oxidation proceeds through the flavin prosthetic group. Synthesis of [2-14C]-3-thia-octanoyl-CoA, followed by isolation of radiolabeled peptide from the modified medium-chain dehydrogenase, showed that inactivation results in labeling the catalytic base, GLU376. Oxidative modification is accompanied by the release of CoASH. A mechanism for inactivation is proposed involving generation of a sulfonium salt which efficiently captures the carboxylate nucleophile.
authors.affiliation: Department of Chemistry and Biochemistry, University of Delaware, Newark 19716, USA.
authors.forename: R A
authors.initials: RA
authors.lastname: Schaller
authors.forename: C
authors.initials: C
authors.lastname: Thorpe
mesh_terms: D000214:Acyl Coenzyme A / Q000737:chemistry / Q000378:metabolism; D042964:Acyl-CoA Dehydrogenase; D044944:Acyl-CoA Dehydrogenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000345:Affinity Labels; D000595:Amino Acid Sequence; D001665:Binding Sites; D003065:Coenzyme A / Q000032:analysis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004952:Esters / Q000378:metabolism; D005296:Ferrous Compounds / Q000494:pharmacology; D005182:Flavin-Adenine Dinucleotide / Q000031:analogs & derivatives / Q000378:metabolism; D005420:Flavoproteins / Q000187:drug effects / Q000378:metabolism*; D008024:Ligands; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry; D017421:Sequence Analysis; D013053:Spectrophotometry; D013438:Sulfhydryl Compounds / Q000378:metabolism
chemical_list: C097160:3-thia-octanoyl-coenzyme A; D000214:Acyl Coenzyme A; D000345:Affinity Labels; D004791:Enzyme Inhibitors; D004952:Esters; D005296:Ferrous Compounds; D005420:Flavoproteins; D008024:Ligands; D010446:Peptide Fragments; D013438:Sulfhydryl Compounds; C064804:ferrocenium; D005182:Flavin-Adenine Dinucleotide; C017123:8-chloro-FAD; D044944:Acyl-CoA Dehydrogenases; D042964:Acyl-CoA Dehydrogenase; D003065:Coenzyme A