title: Chemistry of gene silencing: the mechanism of NAD+-dependent deacetylation reactions.
abstract: The Sir2 enzyme family is responsible for a newly classified chemical reaction, NAD(+)-dependent protein deacetylation. New peptide substrates, the reaction mechanism, and the products of the acetyl transfer to NAD(+) are described for SIR2. The final products of SIR2 reactions are the deacetylated peptide and the 2' and 3' regioisomers of O-acetyl ADP ribose (AADPR), formed through an alpha-1'-acetyl ADP ribose intermediate and intramolecular transesterification reactions (2' --> 3'). The regioisomers, their anomeric forms, the interconversion rates, and the reaction equilibria were characterized by NMR, HPLC, 18O exchange, and MS methods. The mechanism of acetyl transfer to NAD(+) includes (1) ADP ribosylation of the peptide acyl oxygen to form a high-energy O-alkyl amidate intermediate, (2) attack of the 2'-OH group on the amidate to form a 1',2'-acyloxonium species, (3) hydrolysis to 2'-AADPR by the attack of water on the carbonyl carbon, and (4) an SIR2-independent transesterification equilibrating the 2'- and 3'-AADPRs. This mechanism is unprecedented in ADP-ribosyl transferase enzymology. The 2'- and 3'-AADPR products are candidate molecules for SIR2-initiated signaling pathways.
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA.
authors.forename: A A
authors.initials: AA
authors.lastname: Sauve
authors.forename: I
authors.initials: I
authors.lastname: Celic
authors.forename: J
authors.initials: J
authors.lastname: Avalos
authors.forename: H
authors.initials: H
authors.lastname: Deng
authors.forename: J D
authors.initials: JD
authors.lastname: Boeke
authors.forename: V L
authors.initials: VL
authors.lastname: Schramm
mesh_terms: D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000138:chemical synthesis* / Q000737:chemistry / Q000378:metabolism; D000595:Amino Acid Sequence; D001089:Arabinose / Q000737:chemistry; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D017666:Deuterium Oxide / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000737:chemistry; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics; D020868:Gene Silencing; D056572:Histone Deacetylase Inhibitors; D006655:Histone Deacetylases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D007536:Isomerism; D007700:Kinetics; D008969:Molecular Sequence Data; D009243:NAD / Q000737:chemistry* / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D038961:O-Acetyl-ADP-Ribose; D010103:Oxygen Isotopes / Q000378:metabolism; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D056564:Sirtuin 1; D056565:Sirtuin 2; D037761:Sirtuins; D013379:Substrate Specificity; D015534:Trans-Activators / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D019843:Archaeal Proteins; D001426:Bacterial Proteins; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D056572:Histone Deacetylase Inhibitors; D038961:O-Acetyl-ADP-Ribose; D010103:Oxygen Isotopes; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D015534:Trans-Activators; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D001089:Arabinose; C087778:SIR2 protein, S cerevisiae; C447939:SIRT1 protein, human; D056564:Sirtuin 1; D056565:Sirtuin 2; D037761:Sirtuins; D006655:Histone Deacetylases; D017666:Deuterium Oxide