Interaction of 3,4-Dienoyl-CoA Thioesters with Medium Chain Acyl-CoA Dehydrogenase:  Stereochemistry of Inactivation of a Flavoenzyme

Wang, Wenzhong; Fu, Zuoling; Zhou, Jian-Zhong; Kim, Jung‐Ja P.; Thorpe, Colin

doi:10.1021/bi0109818

Cited by 11 publications

(16 citation statements)

References 44 publications

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“…These studies have been extended recently with the synthesis of a series of longer 3,4‐allenes such as R (–)‐3,4‐decadienoyl‐CoA (Fig. 8, compound G) [122]. The R ‐isomer at C‐5 is inhibitory, whereas the S ‐enantiomer is isomerized, apparently directly, to the conjugated 2,4‐diene.…”

Section: Thioesters That Capture the Flavin Prosthetic Groupmentioning

confidence: 99%

“…Somewhat surprisingly, adducts formed from the pantetheine thioesters of these 3,4‐allenes are more kinetically stable than their full‐length CoA counterparts [121,122]. Again, the crystal structures of the ‐pantetheine and ‐N‐acetyl analogs provides a rationalization, with a dramatic change in the coordination environment of the thioester carbonyl between full length and truncated derivatives [122]. Additionally, the kinetic stability of these adducts is strongly influenced by changes in the size of the alkyl substituent at C5 [122].…”

Section: Thioesters That Capture the Flavin Prosthetic Groupmentioning

confidence: 99%

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Acyl‐CoA dehydrogenases

Ghisla

Thorpe

2004

European Journal of Biochemistry

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308

297

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Acyl-CoA dehydrogenases constitute a family of flavoproteins that catalyze the a,b-dehydrogenation of fatty acid acyl-CoA conjugates. While they differ widely in their specificity, they share the same basic chemical mechanism of a,b-dehydrogenation. Medium chain acyl-CoA dehydrogenase is probably the best-studied member of the class and serves as a model for the study of catalytic mechanisms. Based on medium chain acyl-CoA dehydrogenase we discuss the main factors that bring about catalysis, promote specificity and determine the selective transfer of electrons to electron transferring flavoprotein. The mechanism of a,bdehydrogenation is viewed as a process in which the substrate aC-H and bC-H bonds are ruptured concertedly, the first hydrogen being removed by the active center base Glu376-COO -as an H + , the second being transferred as a hydride to the flavin N(5) position. Hereby the pK a of the substrate aC-H is lowered from > 20 to % 8 by the effect of specific hydrogen bonds. Concomitantly, the pK a of Glu376-COO -is also raised to 8-9 due to the decrease in polarity brought about by substrate binding. The kinetic sequence of medium chain acyl-CoA dehydrogenase is rather complex and involves several intermediates. A prominent one is the molecular complex of reduced enzyme with the enoyl-CoA product that is characterized by an intense charge transfer absorption and serves as the point of transfer of electrons to the electron transferring flavoprotein. These views are also discussed in the context of the accompanying paper on the three-dimensional properties of acyl-CoA dehydrogenases.

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Section: Thioesters That Capture the Flavin Prosthetic Groupmentioning

confidence: 99%

Section: Thioesters That Capture the Flavin Prosthetic Groupmentioning

confidence: 99%

Acyl‐CoA dehydrogenases

Ghisla

Thorpe

2004

European Journal of Biochemistry

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308

297

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“…This proton-electron-proton-electron mechanism explains the experimental observations with the radical clock (methylenecyclopropyl)acetyl-CoA that rearranges at the level of the enoxy radical before the second electron transfer (42). However, it has been argued that with normal substrates the reaction appears to be concerted without accumulation of an enolate intermediate able to be oxidized by one-electron transfers (43). Probably, rapid, successive single electron and proton transfers cannot be distinguished from a hydride transfer by the currently available methods.…”

Section: Evolutionary Link With the Mcad Family And Mechanistic Implimentioning

confidence: 87%

Crystal structure of 4-hydroxybutyryl-CoA dehydratase: Radical catalysis involving a [4Fe–4S] cluster and flavin

Martins

Dobbek

Çinkaya

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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Dehydratases catalyze the breakage of a carbonOoxygen bond leading to unsaturated products via the elimination of water. The 1.6-Å resolution crystal structure of 4-hydroxybutyryl-CoA dehydratase from the ␥-aminobutyrate-fermenting Clostridium aminobutyricum represents a new class of dehydratases with an unprecedented active site architecture. A [4Fe-4S] 2؉ cluster, coordinated by three cysteine and one histidine residues, is located 7 Å from the Re-side of a flavin adenine dinucleotide (FAD) moiety. The structure provides insight into the function of these ubiquitous prosthetic groups in the chemically nonfacile, radical-mediated dehydration of 4-hydroxybutyryl-CoA. The substrate can be bound between the [4Fe-4S] 2؉ cluster and the FAD with both cofactors contributing to its radical activation and catalytic conversion. Our results raise interesting questions regarding the mechanism of acyl-CoA dehydrogenases, which are involved in fatty acid oxidation, and address the divergent evolution of the ancestral common gene. Dehydration is a very common reaction in biochemical pathways. More than 100 dehydratases (carbon-oxygen lyases, EC 4.2.-.-) are known [Expert Protein Analysis System (ExPASy), www.expasy.org]. Most of these enzymes catalyze the ␣,␤-elimination of water, during which the ␣-hydrogen (C2 position) to be removed as a proton is activated by an adjacent electron-withdrawing carboxylate, carbonyl, or CoA-thiol ester group, and the hydroxyl group leaves from the ␤-position (C3 position) (1). In anaerobic microorganisms, the absence of the biradical dioxygen allows a rich chemistry of radical reactions involved in the removal of hydrogen atoms from nonactivated positions such as the ␤-or ␥-carbons of 2-, 4-, or 5-hydroxyacylCoA derivatives. These atypical dehydratases contain one or more prosthetic groups such as Fe-S clusters or flavins (2). An example is 4-hydroxybutyryl-CoA dehydratase (4-BUDH) from Clostridium aminobutyricum, which catalyzes both the reversible oxygen-sensitive dehydration of 4-hydroxybutyryl-CoA (Scheme 1a) and the oxygen-insensitive isomerization of vinylacetyl-CoA (Scheme 1c) to crotonyl-CoA (Scheme 1b) (3). The dehydration reaction requires the removal of a hydrogen atom from the least activated C3 position of the butyryl chain (pK a Ϸ 40) and is the mechanistically most demanding step in the fermentation of ␥-aminobutyrate (GABA) to ammonia, acetate, and butyrate by C. aminobutyricum (4). 4-BUDH is active as a homotetramer with up to one [4Fe-4S] 2ϩ cluster and one noncovalently bound flavin adenine dinucleotide (FAD) moiety per 54-kDa subunit. Despite the presence of a [4Fe-4S] 2ϩ cluster and the need of an oxidized FAD for catalysis, the overall reaction occurs with no net redox change (4). These observations have led to the proposal of a radical-based mechanism for the dehydration, where the one-electron oxidation of the enolate of 4-hydroxybutyryl-CoA to the enoxy radical makes the C3-proS-hydrogen (5) acidic enough [pK a ϭ 14 (6)] for deprotonation to a ketyl radical anion (F...

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“…Interest has grown in selective ACADs inhibitors, including MCPA-CoA-like compounds, because of potential therapeutic value in heart disease [46, 47]. MCPA-CoA is the physiological metabolite of hypoglycin A, a naturally occurring toxin in the Ackee plant fruit that peaks just prior to maturity.…”

Section: Discussionmentioning

confidence: 99%

“…Toxic levels of hypoglycin A cause Jamaican vomiting sickness, characterized by severe hypoglycemia [48, 49]. MCPA-CoA is a “suicide” inhibitor that targets IVD, as well as other ACADs, binding covalently to the flavin ring of the essential cofactor [47–50]. Despite the irreversible covalent interaction, MCPA-CoA was unable to effect a change in the absorbance at 447 nm to the same extent as isovaleryl-CoA (only ~72% of the latter) even at very large excess of inhibitor to enzyme ratio.…”

Section: Discussionmentioning

confidence: 99%

Kinetic and spectral properties of isovaleryl-CoA dehydrogenase and interaction with ligands

Mohsen

Vockley

2015

Biochimie

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Isovaleryl-CoA dehydrogenase (IVD) catalyzes the conversion of isovaleryl-CoA to 3-methylcrotonyl-CoA and the transfer of electrons to the electron transfer flavoprotein (ETF). Recombinant human IVD purifies with bound CoA-persulfide. A modified purification protocol was developed to isolate IVD without bound CoA-persulfide and to protect the protein thiols from oxidation. The CoA-persulfide-free IVD specific activity was 112.5 µmol porcine ETF•min−1•mg−1, which was ~20-fold higher than that of its CoA-persulfide bound form. The Km and catalytic efficiency (kcat/Km) for isovaleryl-CoA were 1.0 µM and 4.3 × 106•M−1•sec−1 per monomer, respectively, and its Km for ETF was 2.0 µM. Anaerobic titration of isovaleryl-CoA into an IVD solution resulted in a stable blue complex with increased absorbance at 310 nm, decreased absorbance at 373 and 447 nm, and the appearance of the charge transfer complex band at 584 nm. The apparent dissociation constant (KD app) determined spectrally for isovaleryl-CoA was 0.54 µM. Isovaleryl-CoA, acetoacetyl-CoA, methylenecyclopropylacetyl-CoA, and ETF induced CD spectral changes at the 250–500 nm region while isobutyryl-CoA did not, suggesting conformational changes occur at the flavin ring that are ligand specific. Replacement of the IVD Trp166 with a Phe did not block IVD interaction with ETF, indicating that its indole ring is not essential for electron transfer to ETF. A twelve amino acid synthetic peptide that matches the sequence of the ETF docking peptide competitively inhibited the enzyme reaction when ETF was used as the electron acceptor with a Ki of 1.5 mM.

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Interaction of 3,4-Dienoyl-CoA Thioesters with Medium Chain Acyl-CoA Dehydrogenase: Stereochemistry of Inactivation of a Flavoenzyme

Cited by 11 publications

References 44 publications

Acyl‐CoA dehydrogenases

Acyl‐CoA dehydrogenases

Crystal structure of 4-hydroxybutyryl-CoA dehydratase: Radical catalysis involving a [4Fe–4S] cluster and flavin

Kinetic and spectral properties of isovaleryl-CoA dehydrogenase and interaction with ligands

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