title: Coupling of histone deacetylation to NAD breakdown by the yeast silencing protein Sir2: Evidence for acetyl transfer from substrate to an NAD breakdown product.
abstract: The Saccharomyces cerevisiae silencing protein Sir2 is the founding member of a universally conserved family of proteins that have been shown to possess NAD-dependent histone deacetylation and ADP-ribosylation activities. Here we show that histone deacetylation by Sir2 is coupled to cleavage of the high-energy bond that links the ADP-ribose moiety of NAD to nicotinamide. Analysis of the NAD cleavage products revealed the presence of nicotinamide, ADP-ribose, and a third product that appeared to be related to ADP-ribose. With the use of label transfer experiments, we show that the acetyl group in the histone substrate is transferred to this NAD breakdown product during deacetylation, forming a product that we conclude to be O-acetyl-ADP-ribose. Detection of this species strongly argues for obligate coupling of histone deacetylation to NAD breakdown by Sir2. We propose reaction mechanisms that could account for this coupling via acetyl-ADP-ribose formation. The unprecedented coupling of amide bond cleavage to cleavage of a high-energy bond raises the possibility that NAD breakdown by Sir2 plays an important role in silencing that is independent of its requirement for deacetylation.
authors.affiliation: Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Tanny
authors.forename: D
authors.initials: D
authors.lastname: Moazed
mesh_terms: D000107:Acetylation; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives / Q000096:biosynthesis / Q000378:metabolism; D003067:Coenzymes / Q000502:physiology*; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D015966:Gene Expression Regulation, Fungal; D020868:Gene Silencing; D006655:Histone Deacetylases / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006657:Histones / Q000378:metabolism*; D006868:Hydrolysis; D008954:Models, Biological; D009243:NAD / Q000502:physiology*; D038961:O-Acetyl-ADP-Ribose; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000502:physiology*
chemical_list: D003067:Coenzymes; D005656:Fungal Proteins; D006657:Histones; D038961:O-Acetyl-ADP-Ribose; D011993:Recombinant Fusion Proteins; D038281:Silent Information Regulator Proteins, Saccharomyces cerevisiae; D015534:Trans-Activators; D009243:NAD; D000246:Adenosine Diphosphate Ribose; C087778:SIR2 protein, S cerevisiae; D056565:Sirtuin 2; D037761:Sirtuins; D006655:Histone Deacetylases