@article{Engst1999Mecha-6676,
  year={1999},
  doi={10.1021/bi9815041},
  title={Mechanism of activation of acyl-CoA substrates by medium chain acyl-CoA dehydrogenase : interaction of the thioester carbonyl with the flavin adenine dinucleotide ribityl side chain},
  number={1},
  volume={38},
  issn={0006-2960},
  journal={Biochemistry},
  pages={257--267},
  author={Engst, Stefan and Vock, Petra and Wang, Ming and Kim, Jung-Ja P. and Ghisla, Sandro}
}

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    <dc:contributor>Ghisla, Sandro</dc:contributor>
    <dc:creator>Engst, Stefan</dc:creator>
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    <dc:creator>Ghisla, Sandro</dc:creator>
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    <dcterms:bibliographicCitation>First publ. in: Biochemistry ; 38 (1999), 1. - S. 257-267</dcterms:bibliographicCitation>
    <dc:creator>Wang, Ming</dc:creator>
    <dc:date rdf:datatype="http://www.w3.org/2001/XMLSchema#dateTime">2011-03-24T17:28:17Z</dc:date>
    <dcterms:title>Mechanism of activation of acyl-CoA substrates by medium chain acyl-CoA dehydrogenase : interaction of the thioester carbonyl with the flavin adenine dinucleotide ribityl side chain</dcterms:title>
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    <dc:language>eng</dc:language>
    <dcterms:abstract xml:lang="eng">The flavin adenine dinucleotide (FAD) cofactor of pig kidney medium-chain specific acyl-coenzyme A (CoA) dehydrogenase (MCADH) has been replaced by ribityl-3'-deoxy-FAD and ribityl-2'-deoxy-FAD. 3'-Deoxy-FAD-MCADH has properties very similar to those of native MCADH, indicating that the FAD-ribityl side-chain 3'-OH group does not play any particular role in cofactor binding or catalysis. 2'-Deoxy-FAD-MCADH was characterized using the natural substrate C8CoA as well as various substrate and transition-state analogues. Substrate dehydrogenation in 2'-deoxy-FAD-MCADH is ≈1.5 × 107-fold slower than that of native MCADH, indicating that disruption of the hydrogen bond between 2'-OH and substrate thioester carbonyl leads to a substantial transition-state destabilization equivalent to ≈38 kJ mol-1. The αC-H microscopic pKa of the substrate analogue 3S-C8CoA, which undergoes α-deprotonation on binding to MCADH, is lowered from ≈16 in the free state to ≈11 (±0.5) when bound to 2'-deoxy-FAD-MCADH. This compares with a decrease of the same pKa to ≈5 in the complex with unmodified hwtMCADH, which corresponds to a pK shift of ≈11 pK units, i.e., ≈65 kJ mol-1 [Vock, P., Engst, S., Eder, M., and Ghisla, S. (1998) Biochemistry 37, 1848-1860]. The difference of this effect of ≈6 pK units (≈35 kJ mol-1) between MCADH and 2'-deoxy-FAD-MCADH is taken as the level of stabilization of the substrate carbanionic species caused by the interaction with the FAD-2'-OH. This energetic parameter derived from the kinetic experiments (stabilization of transition state) is in agreement with those obtained from static experiments (lowering of αC-H microscopic pKa of analogue, i.e., stabilization of anionic transition-state analogue). The contributions of the two single H-bonds involved in substrate activation (Glu376amide-N-H and ribityl-2'-OH) thus appear to behave additively toward the total effect. The crystal structures of native pMCADH and of 2'-deoxy-FAD-MCADH complexed with octanoyl-CoA/octenoyl-CoA show unambiguously that the FAD cofactor and the substrate/product bind in an identical fashion, implying that the observed effects are mainly due to (the absence of) the FAD-ribityl-2'-OH hydrogen bond. The large energy associated with the 2'-OH hydrogen bond interaction is interpreted as resulting from the changes in charge and the increased hydrophobicity induced by binding of lipophilic substrate. This is the first example demonstrating the direct involvement of a flavin cofactor side chain in catalysis.</dcterms:abstract>
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    <dcterms:issued>1999</dcterms:issued>
    <dc:contributor>Vock, Petra</dc:contributor>
    <dc:creator>Kim, Jung-Ja P.</dc:creator>
    <dc:contributor>Wang, Ming</dc:contributor>
    <dc:creator>Vock, Petra</dc:creator>
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    <dc:contributor>Kim, Jung-Ja P.</dc:contributor>
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