title: Thermal unfolding of medium-chain acyl-CoA dehydrogenase and iso(3)valeryl-CoA dehydrogenase: study of the effect of genetic defects on enzyme stability.
abstract: Genetic defects affecting acyl-CoA dehydrogenases (ACAD)-key enzymes in the degradation of fatty acids and branched chain amino acids-are increasingly recognized as being more widespread than originally thought. For the medium-chain acyl-CoA dehydrogenase (MCAD), the K304E mutation is the most common genetic defect among Caucasian populations. The effect of substrate or substrate analog binding on the stability of wild-type MCAD and isovaleryl-CoA dehydrogenase (i3VD) and their genetic mutants (K304E- and T168A-MCAD and A282V-i3VD) is examined. Binding to the mutant ACADs is generally approximately 10-fold weaker compared to wild-type proteins. Thermal stability of wt-MCAD (melting point approximately 53.6 degrees C) is significantly higher compared to wt-i3VD ( approximately 49.3 degrees C). With the exception of the A282V-i3VD mutant, a high degree of stabilization (5-11 degrees C) is induced by conversion into the reduced enzyme form complexed with product. The results are discussed based on the 3D-structures of the enzymes, and it is concluded that in the case of K304E-MCAD thermal stability as such is not a major contribution to the clinical phenotype. With the T168A-MCAD and A282V-i3VD mutants, however, the diminished thermal stability and minor stabilization by ligands must be regarded as an important factor contributing to the manifestation of the disease.
authors.affiliation: Department of Biology, University of Konstanz, P.O. Box 5560-M644, D-78457 Konstanz, Germany.
authors.forename: Ibrahim
authors.initials: I
authors.lastname: Nasser
authors.forename: Al-Walid
authors.initials: AW
authors.lastname: Mohsen
authors.forename: Ilian
authors.initials: I
authors.lastname: Jelesarov
authors.forename: Jerry
authors.initials: J
authors.lastname: Vockley
authors.forename: Peter
authors.initials: P
authors.lastname: Macheroux
authors.forename: Sandro
authors.initials: S
authors.lastname: Ghisla
mesh_terms: D042964:Acyl-CoA Dehydrogenase / Q000737:chemistry* / Q000235:genetics; D004795:Enzyme Stability; D006358:Hot Temperature; D050769:Isovaleryl-CoA Dehydrogenase; D008958:Models, Molecular; D009154:Mutation; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000737:chemistry* / Q000235:genetics; D011485:Protein Binding; D017510:Protein Folding
chemical_list: D044925:Oxidoreductases Acting on CH-CH Group Donors; D050769:Isovaleryl-CoA Dehydrogenase; D042964:Acyl-CoA Dehydrogenase