Human acyl-CoA dehydrogenase-9 plays a novel role in the mitochondrial beta-oxidation of unsaturated fatty acids.
Unsaturated fatty acids play an important role in the prevention of human diseases such as diabetes, obesity, cancer, and neurodegeneration. However, their oxidation in vivo by acyl-CoA dehydrogenases (ACADs) that catalyze the first step of each cycle of mitochondrial fatty acid beta-oxidation is not entirely understood. Recently, a novel ACAD (ACAD-9) of unknown function that is highly homologous to human very-long-chain acyl-CoA dehydrogenase was identified by large-scale random sequencing. To characterize its enzymatic role, we have expressed ACAD-9 in Escherichia coli, purified it, and determined its pattern of substrate utilization. The N terminus of the mature form of the enzyme was identified by in vitro mitochondrial import studies of precursor protein. A 37-amino acid leader peptide was cleaved sequentially by two mitochondrial peptidases to yield a predicted molecular mass of 65 kDa for the mature subunit. Submitochondrial fractionation studies found native ACAD-9 to be associated with the mitochondrial membrane. Gel filtration analysis indicated that, like very-long-chain acyl-CoA dehydrogenase, ACAD-9 is a dimer, in contrast to the other known ACADs, which are tetramers. Purified mature ACAD-9 had maximal activity with long-chain unsaturated acyl-CoAs as substrates (C16:1-, C18:1-, C18:2-, C22:6-CoA). These results suggest a previously unrecognized role for ACAD-9 in the mitochondrial beta-oxidation of long-chain unsaturated fatty acids. Because of the substrate specificity and abundance of ACAD-9 in brain, we speculate that it may play a role in the turnover of lipid membrane unsaturated fatty acids that are essential for membrane integrity and structure.
Ensenauer
Regina
R
Department of Medical Genetics, Pediatric and Adolescent Medicine, Mayo Clinic College of Medicine, Rochester, Minnesota 55905, USA.
He
Miao
M
Willard
Jan-Marie
JM
Goetzman
Eric S
ES
Corydon
Thomas J
TJ
Vandahl
Brian B
BB
Mohsen
Al-Walid
AW
Isaya
Grazia
G
Vockley
Jerry
J
D044942:Acyl-CoA Dehydrogenase, Long-Chain / Q000378:metabolism / Q000502:physiology*; D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D008928:Mitochondria / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D016296:Mutagenesis; D010100:Oxygen / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D014158:Transcription, Genetic
D005227:Fatty Acids; D008055:Lipids; D010455:Peptides; D011994:Recombinant Proteins; D044942:Acyl-CoA Dehydrogenase, Long-Chain; C467805:ACADVL protein, human; D010100:Oxygen