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| | ==Crystal structure of the yeast nicotinamidase Pnc1p bound to the inhibitor nicotinaldehyde== | | ==Crystal structure of the yeast nicotinamidase Pnc1p bound to the inhibitor nicotinaldehyde== |
| - | <StructureSection load='3v8e' size='340' side='right' caption='[[3v8e]], [[Resolution|resolution]] 2.71Å' scene=''> | + | <StructureSection load='3v8e' size='340' side='right'caption='[[3v8e]], [[Resolution|resolution]] 2.71Å' scene=''> |
| | == Structural highlights == | | == Structural highlights == |
| - | <table><tr><td colspan='2'>[[3v8e]] is a 7 chain structure with sequence from [http://en.wikipedia.org/wiki/Saccharomyces_cerevisiae Saccharomyces cerevisiae]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=3V8E OCA]. For a <b>guided tour on the structure components</b> use [http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=3V8E FirstGlance]. <br> | + | <table><tr><td colspan='2'>[[3v8e]] is a 7 chain structure with sequence from [https://en.wikipedia.org/wiki/Saccharomyces_cerevisiae_S288C Saccharomyces cerevisiae S288C]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=3V8E OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=3V8E FirstGlance]. <br> |
| - | </td></tr><tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat"><scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr> | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.71Å</td></tr> |
| - | <tr id='NonStdRes'><td class="sblockLbl"><b>[[Non-Standard_Residue|NonStd Res:]]</b></td><td class="sblockDat"><scene name='pdbligand=JJJ:S-(PYRIDIN-3-YLCARBONYL)-L-CYSTEINE'>JJJ</scene></td></tr> | + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=JJJ:S-(PYRIDIN-3-YLCARBONYL)-L-CYSTEINE'>JJJ</scene>, <scene name='pdbligand=MG:MAGNESIUM+ION'>MG</scene>, <scene name='pdbligand=ZN:ZINC+ION'>ZN</scene></td></tr> |
| - | <tr id='related'><td class="sblockLbl"><b>[[Related_structure|Related:]]</b></td><td class="sblockDat">[[2h0r|2h0r]]</td></tr>
| + | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[https://proteopedia.org/fgij/fg.htm?mol=3v8e FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=3v8e OCA], [https://pdbe.org/3v8e PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=3v8e RCSB], [https://www.ebi.ac.uk/pdbsum/3v8e PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=3v8e ProSAT]</span></td></tr> |
| - | <tr id='gene'><td class="sblockLbl"><b>[[Gene|Gene:]]</b></td><td class="sblockDat">PNC1, YGL037C ([http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&srchmode=5&id=4932 Saccharomyces cerevisiae])</td></tr>
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| - | <tr id='activity'><td class="sblockLbl"><b>Activity:</b></td><td class="sblockDat"><span class='plainlinks'>[http://en.wikipedia.org/wiki/Nicotinamidase Nicotinamidase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=3.5.1.19 3.5.1.19] </span></td></tr>
| + | |
| - | <tr id='resources'><td class="sblockLbl"><b>Resources:</b></td><td class="sblockDat"><span class='plainlinks'>[http://oca.weizmann.ac.il/oca-docs/fgij/fg.htm?mol=3v8e FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=3v8e OCA], [http://www.rcsb.org/pdb/explore.do?structureId=3v8e RCSB], [http://www.ebi.ac.uk/pdbsum/3v8e PDBsum]</span></td></tr> | + | |
| | </table> | | </table> |
| | == Function == | | == Function == |
| - | [[http://www.uniprot.org/uniprot/PNC1_YEAST PNC1_YEAST]] Catalyzes the deamidation of nicotinamide, an early step in the NAD(+) salvage pathway. Positively regulates SIR2-mediated silencing and longevity by preventing the accumulation of intracellular nicotinamide, an inhibitor of SIR2, during times of stress. Acts also on nicotinyl hydroxamate.<ref>PMID:11816029</ref> <ref>PMID:12736687</ref> <ref>PMID:14729974</ref> | + | [https://www.uniprot.org/uniprot/PNC1_YEAST PNC1_YEAST] Catalyzes the deamidation of nicotinamide, an early step in the NAD(+) salvage pathway. Positively regulates SIR2-mediated silencing and longevity by preventing the accumulation of intracellular nicotinamide, an inhibitor of SIR2, during times of stress. Acts also on nicotinyl hydroxamate.<ref>PMID:11816029</ref> <ref>PMID:12736687</ref> <ref>PMID:14729974</ref> |
| | <div style="background-color:#fffaf0;"> | | <div style="background-color:#fffaf0;"> |
| | == Publication Abstract from PubMed == | | == Publication Abstract from PubMed == |
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| | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> | | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| | </div> | | </div> |
| | + | <div class="pdbe-citations 3v8e" style="background-color:#fffaf0;"></div> |
| | == References == | | == References == |
| | <references/> | | <references/> |
| | __TOC__ | | __TOC__ |
| | </StructureSection> | | </StructureSection> |
| - | [[Category: Nicotinamidase]] | + | [[Category: Large Structures]] |
| - | [[Category: Saccharomyces cerevisiae]] | + | [[Category: Saccharomyces cerevisiae S288C]] |
| - | [[Category: Denu, J M]] | + | [[Category: Denu JM]] |
| - | [[Category: Hoadley, K A]] | + | [[Category: Hoadley KA]] |
| - | [[Category: Keck, J L]] | + | [[Category: Keck JL]] |
| - | [[Category: Smith, B C]] | + | [[Category: Smith BC]] |
| - | [[Category: Hydrolase]]
| + | |
| Structural highlights
Function
PNC1_YEAST Catalyzes the deamidation of nicotinamide, an early step in the NAD(+) salvage pathway. Positively regulates SIR2-mediated silencing and longevity by preventing the accumulation of intracellular nicotinamide, an inhibitor of SIR2, during times of stress. Acts also on nicotinyl hydroxamate.[1] [2] [3]
Publication Abstract from PubMed
Nicotinamidases catalyze the hydrolysis of nicotinamide to nicotinic acid and ammonia. Nicotinamidases are absent in mammals but function in NAD(+) salvage in many bacteria, yeast, plants, protozoa, and metazoans. We have performed structural and kinetic investigations of the nicotinamidase from Saccharomyces cerevisiae (Pnc1). Steady-state product inhibitor analysis revealed an irreversible reaction in which ammonia is the first product released, followed by nicotinic acid. A series of nicotinamide analogues acting as inhibitors or substrates were examined, revealing that the nicotinamide carbonyl oxygen and ring nitrogen are critical for binding and reactivity. X-ray structural analysis revealed a covalent adduct between nicotinaldehyde and Cys167 of Pnc1 and coordination of the nicotinamide ring nitrogen to the active-site zinc ion. Using this structure as a guide, the function of several residues was probed via mutagenesis and primary (15)N and (13)C kinetic isotope effects (KIEs) on V/K for amide bond hydrolysis. The KIE values of almost all variants were increased, indicating that C-N bond cleavage is at least partially rate limiting; however, a decreased KIE for D51N was indicative of a stronger commitment to catalysis. In addition, KIE values using slower alternate substrates indicated that C-N bond cleavage is at least partially rate limiting with nicotinamide to highly rate limiting with thionicotinamide. A detailed mechanism involving nucleophilic attack of Cys167, followed by elimination of ammonia and then hydrolysis to liberate nicotinic acid, is discussed. These results will aid in the design of mechanism-based inhibitors to target pathogens that rely on nicotinamidase activity.
Structural and Kinetic Isotope Effect Studies of Nicotinamidase (Pnc1) from Saccharomyces cerevisiae.,Smith BC, Anderson MA, Hoadley KA, Keck JL, Cleland WW, Denu JM Biochemistry. 2012 Jan 10;51(1):243-56. Epub 2011 Dec 29. PMID:22229411[4]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
References
- ↑ Ghislain M, Talla E, Francois JM. Identification and functional analysis of the Saccharomyces cerevisiae nicotinamidase gene, PNC1. Yeast. 2002 Feb;19(3):215-24. PMID:11816029 doi:http://dx.doi.org/10.1002/yea.810
- ↑ Anderson RM, Bitterman KJ, Wood JG, Medvedik O, Sinclair DA. Nicotinamide and PNC1 govern lifespan extension by calorie restriction in Saccharomyces cerevisiae. Nature. 2003 May 8;423(6936):181-5. PMID:12736687 doi:http://dx.doi.org/10.1038/nature01578
- ↑ Gallo CM, Smith DL Jr, Smith JS. Nicotinamide clearance by Pnc1 directly regulates Sir2-mediated silencing and longevity. Mol Cell Biol. 2004 Feb;24(3):1301-12. PMID:14729974
- ↑ Smith BC, Anderson MA, Hoadley KA, Keck JL, Cleland WW, Denu JM. Structural and Kinetic Isotope Effect Studies of Nicotinamidase (Pnc1) from Saccharomyces cerevisiae. Biochemistry. 2012 Jan 10;51(1):243-56. Epub 2011 Dec 29. PMID:22229411 doi:10.1021/bi2015508
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