The reductive half-reaction in Acyl-CoA dehydrogenase from pig kidney: studies with thiaoctanoyl-CoA and oxaoctanoyl-CoA analogues.
Thia- and oxaoctanoyl-CoA derivatives (substituted at the C-3 and C-4 positions) have been synthesized to prove the reductive half-reaction in the medium-chain acyl-CoA dehydrogenase from pig kidney. 3-Thiaoctanoyl-CoA binds to this flavoenzyme, forming an intense, stable, long-wavelength band (at 804 nm; extinction coefficient = 8.7 mM-1 cm-1 at pH 7.6). The intensity of this band increases about 20% from pH 6.0 to pH 8.8. This long-wavelength species probably represents a charge-transfer complex between bound acyl enolate as the donor and oxidized flavin adenine dinucleotide as the acceptor. Thus, the enzyme catalyzes alpha-proton exchange, and no long-wavelength bands are seen with 3-thiaoctyl-CoA (where the carbonyl moiety is replaced by a methylene group). 3-Oxaoctanoyl-CoA binds comparatively weakly to the dehydrogenase, with a long-wavelength band at 780 nm which is both less intense and less stable than the corresponding thia analogue. These data suggest that the enzyme can accomplish alpha-proton abstraction from certain weakly acidic acyl-CoA derivatives, without concerted transfer of a hydride equivalent to the flavin. 4-Thiaoctanoyl-CoA is dehydrogenated in the standard assay 1.5-fold faster than octanoyl-CoA. Titrations of the medium-chain dehydrogenase with the 4-thia derivative resemble those obtained with octanoyl-CoA, except for the contribution of the strongly absorbing 4-thia-trans-2-octenoyl-CoA product. The corresponding 4-oxa analogue is a much poorer substrate (10% of the rate shown by octanoyl-CoA) but again effects substantially complete reduction of the flavin chromophore in the dehydrogenase.(ABSTRACT TRUNCATED AT 250 WORDS)
Lau
S M
SM
Department of Chemistry and Biochemistry, University of Delaware, Newark 19716.
Brantley
R K
RK
Thorpe
C
C
D000214:Acyl Coenzyme A / Q000378:metabolism*; D044944:Acyl-CoA Dehydrogenases / Q000378:metabolism*; D000693:Anaerobiosis; D000818:Animals; D007668:Kidney / Q000201:enzymology*; D013329:Structure-Activity Relationship; D013552:Swine
D000214:Acyl Coenzyme A; D044944:Acyl-CoA Dehydrogenases