Publications

• Identification of the active site catalytic residue in human isovaleryl-CoA dehydrogenase.

  Mohsen A.W., Vockley J.

  Isovaleryl-CoA dehydrogenase (IVD) is a homotetrameric flavoenzyme which catalyzes the conversion of isovaleryl-CoA to 3-methylcrotonyl-CoA. E376 of pig medium chain acyl-CoA dehydrogenase (MCAD), a homologous enzyme, has been identified as the active site catalytic residue. Amino acid sequence al ... >> More

  Isovaleryl-CoA dehydrogenase (IVD) is a homotetrameric flavoenzyme which catalyzes the conversion of isovaleryl-CoA to 3-methylcrotonyl-CoA. E376 of pig medium chain acyl-CoA dehydrogenase (MCAD), a homologous enzyme, has been identified as the active site catalytic residue. Amino acid sequence alignment shows that A375 is the corresponding residue in human IVD. Using the atomic coordinates determined for MCAD, molecular modeling suggests that E254 is the substituting catalytic residue in IVD. To substantiate the importance of this residue for enzyme function, cDNAs for the wild-type human IVD and E254G, E254D, E254Q, and E254G/A375E mutant IVDs were constructed and cloned into a prokaryotic expression vector. The proteins were synthesized in Escherichia coli and purified, and their properties were examined. The catalytic activity of the recombinant wild-type IVD was the highest in the presence of isovaleryl-CoA, and its UV/visible light spectrum in the presence of isovaleryl-CoA showed quenching of its characteristic absorption in the 445-nm region and appearance of absorption at 600 nm. The E254G and E254Q mutant IVDs had no detectable enzymatic activity, and isovaleryl-CoA did not induce quenching of the absorption in the 445-nm region or the appearance of absorption at 600 nm. The E254D mutant IVD had residual activity for isovaleryl-CoA, and its spectrum was altered compared to that of the wild type. The E254G/A375E mutant IVD exhibited catalytic activity toward isovaleryl-CoA, and its spectrum in the absence or presence of the substrate was similar to that of the wild-type IVD.(ABSTRACT TRUNCATED AT 250 WORDS) << Less

  Biochemistry 34:10146-10152(1995) [PubMed] [EuropePMC]

  This publication is cited by 2 other entries.

• Medium-long-chain chimeric human Acyl-CoA dehydrogenase: medium-chain enzyme with the active center base arrangement of long-chain Acyl-CoA dehydrogenase.

  Nandy A., Kieweg V., Kraeutle F.G., Vock P., Kuechler B., Bross P., Kim J.J., Rasched I., Ghisla S.

  The catalytically essential glutamate residue that initiates catalysis by abstracting the substrate alpha-hydrogen as H+ is located at position 376 (mature MCADH numbering) on loop JK in medium chain acyl-CoA dehydrogenase (MCADH). In long chain acyl-CoA dehydrogenase (LCADH) and isovaleryl-CoA de ... >> More

  The catalytically essential glutamate residue that initiates catalysis by abstracting the substrate alpha-hydrogen as H+ is located at position 376 (mature MCADH numbering) on loop JK in medium chain acyl-CoA dehydrogenase (MCADH). In long chain acyl-CoA dehydrogenase (LCADH) and isovaleryl-CoA dehydrogenase (IVDH), the corresponding Glu carrying out the same function is placed at position 255 on the adjacent helix G. These glutamates thus act on substrate approaching from two opposite regions at the active center. We have implemented the topology of LCADH in MCADH by carrying out the two mutations Glu376Gly and Thr255Glu. The resulting chimeric enzyme, "medium-/long" chain acyl-CoA dehydrogenase (MLCADH) has approximately 20% of the activity of MCADH and approximately 25% that of LCADH with its best substrates octanoyl-CoA and dodecanoyl-CoA, respectively. MLCADH exhibits an enhanced rate of reoxidation with oxygen, however, with a much narrower substrate chain length specificity that peaks with dodecanoyl-CoA. This is the same maximum as that of LCADH and is thus significantly shifted from that of native MCADH (hexanoyl/octanoyl-CoA). The putative, common ancestor of LCADH and IVDH has two Glu residues, one each at positions 255 and 376. The corresponding MCADH mutant, Thr255Glu (glu/glu-MCADH), is as active as MCADH with octanoyl-CoA; its activity/chain length profile is, however, much narrower. The topology of the Glu as H+ abstracting base seems an important factor in determining chain length specificity and reactivity in acyl-CoA dehydrogenases. The mechanisms underlying these effects are discussed in view of the three-dimensional structure of MLCADH, which is presented in the accompanying paper [Lee et al. (1996) Biochemistry 35, 12412-12420]. << Less

  Biochemistry 35:12402-12411(1996) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.

• Identification and characterization of new long chain acyl-CoA dehydrogenases.

  He M., Pei Z., Mohsen A.W., Watkins P., Murdoch G., Van Veldhoven P.P., Ensenauer R., Vockley J.

  Long-chain fatty acids are an important source of energy in muscle and heart where the acyl-CoA dehydrogenases (ACADs) participate in consecutive cycles of β-oxidation to generate acetyl-CoA and reducing equivalents for generating energy. However, the role of long-chain fatty acid oxidation in the ... >> More

  Long-chain fatty acids are an important source of energy in muscle and heart where the acyl-CoA dehydrogenases (ACADs) participate in consecutive cycles of β-oxidation to generate acetyl-CoA and reducing equivalents for generating energy. However, the role of long-chain fatty acid oxidation in the brain and other tissues that do not rely on fat for energy is poorly understood. Here we characterize two new ACADs, ACAD10 and ACAD11, both with significant expression in human brain. ACAD11 utilizes substrates with primary carbon chain lengths between 20 and 26, with optimal activity towards C22CoA. The combination of ACAD11 with the newly characterized ACAD9 accommodates the full spectrum of long chain fatty acid substrates presented to mitochondrial β-oxidation in human cerebellum. ACAD10 has significant activity towards the branched-chain substrates R and S, 2 methyl-C15-CoA and is highly expressed in fetal but not adult brain. This pattern of expression is similar to that of LCAD, another ACAD previously shown to be involved in long branched chain fatty acid metabolism. Interestingly, the ACADs in human cerebellum were found to have restricted cellular distribution. ACAD9 was most highly expressed in the granular layer, ACAD11 in the white matter, and MCAD in the molecular layer and axons of specific neurons. This compartmentalization of ACADs in the human central nerve system suggests that β-oxidation in cerebellum participates in different functions other than generating energy, for example, the synthesis and/or degradation of unique cellular lipids and catabolism of aromatic amino acids, compounds that are vital to neuronal function. << Less

  Mol. Genet. Metab. 102:418-429(2011) [PubMed] [EuropePMC]

  This publication is cited by 13 other entries.

• Isolation and expression of a cDNA encoding the precursor for a novel member (ACADSB) of the acyl-CoA dehydrogenase gene family.

  Rozen R., Vockley J., Zhou L., Milos R., Willard J., Fu K., Vicanek C., Low-Nang L., Torban E., Fournier B.

  The acyl-CoA dehydrogenases (ACDs) are a family of mitochondrial enzymes that oxidize straight chain or branched chain acyl-CoAs in the metabolism of fatty acids or branched chain amino acids. Deficiencies in members of this gene family are important causes of human disease. A cDNA encoding the hu ... >> More

  The acyl-CoA dehydrogenases (ACDs) are a family of mitochondrial enzymes that oxidize straight chain or branched chain acyl-CoAs in the metabolism of fatty acids or branched chain amino acids. Deficiencies in members of this gene family are important causes of human disease. A cDNA encoding the human precursor for a novel member (gene symbol ACADSB) of the ACD gene family has been isolated and characterized. The open reading frame of 1.3 kb encodes a precursor protein of 431 amino acids, which is processed in vitro to yield a mature protein of 399 amino acids. The cDNA has significant sequence similarity to other members of the acyl-CoA dehydrogenase family, with the greatest homology (38%) to the short chain acyl-CoA dehydrogenase. The cDNA was expressed in eukaryotic (COS) and prokaryotic (Escherichia coli) cells, producing a protein of the expected size, with activity toward the short branched chain acyl-CoA derivatives ((S)-2-methylbutyryl-CoA, isobutyryl-CoA, and 2-methylhexanoyl-CoA), as well as toward the short straight chain acyl-CoAs (butyryl-CoA and hexanoyl-CoA). << Less

  Genomics 24:280-287(1994) [PubMed] [EuropePMC]

  This publication is cited by 6 other entries.