Substrate activating mechanism of short-chain acyl-CoA, medium-chain acyl-CoA, long-chain acyl-CoA, and isovaleryl-CoA dehydrogenases from bovine liver: a resonance Raman study on the 3-ketoacyl-CoA complexes.
We have shown previously that acetoacetyl-CoA bound to medium-chain acyl-CoA dehydrogenase from pig kidney is transformed into an enolate form, O = C(3)-C(2)H = C(1)-O-, and that the interaction between the C(4a) = N(5) moiety of flavin and the O = C(3)-C(2)H = C(1)-O- moiety of acetoacetyl-CoA is important for the charge-transfer interaction [Nishina, Y. et al. (1992) J. Biochem. 111, 699-706]. In this study, we examined four kinds of acyl-CoA dehydrogenases [short-chain acyl-CoA (SCAD), medium-chain acyl-CoA (MCAD), long-chain acyl-CoA (LCAD), and isovaleryl-CoA (IVD) dehydrogenases] from bovine liver. The Raman spectra of non-labeled and isotopically labeled acetoacetyl-CoA in keto-form revealed that the 1,716-cm-1 and 1,650-cm-1 bands were derived from the C(3) = O and the C(1) = O stretching mode, respectively. In the charge-transfer complexes of acetoacetyl-CoA with the four kinds of dehydrogenases, the resonance Raman (RR) bands corresponding to the C(3) = O and the C(1) = O of acetoacetyl-CoA were observed at around 1,643-1,622 and 1,506-1,476 cm-1, respectively, indicating that acetoacetyl-CoA was transformed into the enolate form as the result of the complexation with the enzymes. Further, in RR spectra with excitation at 632.8 nm, within the charge-transfer band of the complexes of acetoacetyl-CoA with the four acyl-CoA dehydrogenases, both bands associated with the C(4a) = N(5) moiety of oxidized flavin and the O = C(3)-C(2)H = C(1)-O- moiety of acetoacetyl-CoA were enhanced, but the benzene portion of oxidized flavin was not. These results indicate that the substrate activating mechanism is common to all four kinds of dehydrogenases, i.e., the interaction between the C(1) = O of acetoacetyl-CoA and the positively polarized atoms of the enzymes located in close proximity to the oxygen atom of C(1) = O is important, and the C(4a) = N(5) moiety of flavin participates in the interaction. Some kinds of 3-ketoacyl-CoAs were tested instead of acetoacetyl-CoA and essentially similar results were obtained. The positions of the bands derived from the C(1)-O- moiety of 3-ketoacyl-CoAs were different by ca. 30 cm-1 in two groups, i.e., ca. 1,475 cm-1 for SCAD and MCAD and ca. 1,505 cm-1 for LCAD and IVD, that is, RR spectra can classify the four dehydrogenases into two groups.
Hazekawa
I
I
Department of Metabolic Medicine, Kumamoto University School of Medicine, Honjo.
Nishina
Y
Y
Sato
K
K
Shichiri
M
M
Shiga
K
K
D042964:Acyl-CoA Dehydrogenase; D044942:Acyl-CoA Dehydrogenase, Long-Chain / Q000737:chemistry*; D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D006863:Hydrogen-Ion Concentration; D050769:Isovaleryl-CoA Dehydrogenase; D008099:Liver / Q000201:enzymology*; D015394:Molecular Structure; D010088:Oxidoreductases / Q000737:chemistry*; D044925:Oxidoreductases Acting on CH-CH Group Donors; D012996:Solutions; D013059:Spectrum Analysis, Raman
D012996:Solutions; D010088:Oxidoreductases; D000429:Alcohol Oxidoreductases; C023744:acetoacetyl-CoA reductase; D044925:Oxidoreductases Acting on CH-CH Group Donors; D050769:Isovaleryl-CoA Dehydrogenase; D042964:Acyl-CoA Dehydrogenase; D044942:Acyl-CoA Dehydrogenase, Long-Chain