title: Sirtuin 3 (SIRT3) protein regulates long-chain acyl-CoA dehydrogenase by deacetylating conserved lysines near the active site.
abstract: Long-chain acyl-CoA dehydrogenase (LCAD) is a key mitochondrial fatty acid oxidation enzyme. We previously demonstrated increased LCAD lysine acetylation in SIRT3 knockout mice concomitant with reduced LCAD activity and reduced fatty acid oxidation. To study the effects of acetylation on LCAD and determine sirtuin 3 (SIRT3) target sites, we chemically acetylated recombinant LCAD. Acetylation impeded substrate binding and reduced catalytic efficiency. Deacetylation with recombinant SIRT3 partially restored activity. Residues Lys-318 and Lys-322 were identified as SIRT3-targeted lysines. Arginine substitutions at Lys-318 and Lys-322 prevented the acetylation-induced activity loss. Lys-318 and Lys-322 flank residues Arg-317 and Phe-320, which are conserved among all acyl-CoA dehydrogenases and coordinate the enzyme-bound FAD cofactor in the active site. We propose that acetylation at Lys-318/Lys-322 causes a conformational change which reduces hydride transfer from substrate to FAD. Medium-chain acyl-CoA dehydrogenase and acyl-CoA dehydrogenase 9, two related enzymes with lysines at positions equivalent to Lys-318/Lys-322, were also efficiently deacetylated by SIRT3 following chemical acetylation. These results suggest that acetylation/deacetylation at Lys-318/Lys-322 is a mode of regulating fatty acid oxidation. The same mechanism may regulate other acyl-CoA dehydrogenases.
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Sivakama S
authors.initials: SS
authors.lastname: Bharathi
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Yuxun
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Al-Walid
authors.initials: AW
authors.lastname: Mohsen
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Radha
authors.initials: R
authors.lastname: Uppala
authors.affiliation: Genomics and Proteomics Core Facility, University of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Manimalha
authors.initials: M
authors.lastname: Balasubramani
authors.affiliation: Genomics and Proteomics Core Facility, University of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Emanuel
authors.initials: E
authors.lastname: Schreiber
authors.affiliation: Genomics and Proteomics Core Facility, University of Pittsburgh, Pittsburgh, Pennsylvania 15224.
authors.forename: Guy
authors.initials: G
authors.lastname: Uechi
authors.affiliation: Department of Human Genetics, University of Pittsburgh, Graduate School of Public Health, Pittsburgh, Pennsylvania 15224.
authors.forename: Megan E
authors.initials: ME
authors.lastname: Beck
authors.affiliation: Buck Institute for Research on Aging, Novato, California 94945.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Rardin
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224; Department of Human Genetics, University of Pittsburgh, Graduate School of Public Health, Pittsburgh, Pennsylvania 15224.
authors.forename: Jerry
authors.initials: J
authors.lastname: Vockley
authors.affiliation: Gladstone Institutes and University of California, San Francisco, California 94158.
authors.forename: Eric
authors.initials: E
authors.lastname: Verdin
authors.affiliation: Buck Institute for Research on Aging, Novato, California 94945.
authors.forename: Bradford W
authors.initials: BW
authors.lastname: Gibson
authors.affiliation: Sarah W. Stedman Nutrition and Metabolism Center Duke University, Medical Center, Durham, North Carolina 27704.
authors.forename: Matthew D
authors.initials: MD
authors.lastname: Hirschey
authors.affiliation: Department of Pediatrics, University of Pittsburgh School of Medicine, University of Pittsburgh, Children's Hospital of Pittsburgh, Pittsburgh, Pennsylvania 15224; Department of Human Genetics, University of Pittsburgh, Graduate School of Public Health, Pittsburgh, Pennsylvania 15224. Electronic address: eric.goetzman@chp.edu.
authors.forename: Eric S
authors.initials: ES
authors.lastname: Goetzman
mesh_terms: D000107:Acetylation; D044942:Acyl-CoA Dehydrogenase, Long-Chain; D000818:Animals; D020134:Catalytic Domain / Q000502:physiology; D005227:Fatty Acids / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008930:Mitochondria, Liver / Q000201:enzymology* / Q000235:genetics; D010084:Oxidation-Reduction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D056566:Sirtuin 3 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D005227:Fatty Acids; D011994:Recombinant Proteins; C525046:Sirt3 protein, mouse; D005182:Flavin-Adenine Dinucleotide; D044942:Acyl-CoA Dehydrogenase, Long-Chain; C466093:SIRT3 protein, human; D056566:Sirtuin 3
keywords: Acyl-CoA Dehydrogenase; Electron Transfer; Enzyme Catalysis; FAD; Fatty Acid Oxidation; Lysine Acetylation; Mitochondria; Posttranslational Modification; Sirtuins