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| - | [[Image:1d6h.jpg|left|200px]]<br /><applet load="1d6h" size="450" color="white" frame="true" align="right" spinBox="true" | |
| - | caption="1d6h, resolution 2.15Å" /> | |
| - | '''CHALONE SYNTHASE (N336A MUTANT COMPLEXED WITH COA)'''<br /> | |
| | | | |
| - | ==Overview== | + | ==CHALONE SYNTHASE (N336A MUTANT COMPLEXED WITH COA)== |
| - | Chalcone synthase (CHS) catalyzes formation of the phenylpropanoid, chalcone from one p-coumaroyl-CoA and three malonyl-coenzyme A (CoA), thioesters. The three-dimensional structure of CHS [Ferrer, J.-L., Jez, J., M., Bowman, M. E., Dixon, R. A., and Noel, J. P. (1999) Nat. Struct. Biol., 6, 775-784] suggests that four residues (Cys164, Phe215, His303, and, Asn336) participate in the multiple decarboxylation and condensation, reactions catalyzed by this enzyme. Here, we functionally characterize 16, point mutants of these residues for chalcone production, malonyl-CoA, decarboxylation, and the ability to bind CoA and acetyl-CoA. Our results, confirm Cys164's role as the active-site nucleophile in polyketide, formation and elucidate the importance of His303 and Asn336 in the, malonyl-CoA decarboxylation reaction. We suggest that Phe215 may help, orient substrates at the active site during elongation of the polyketide, intermediate. To better understand the structure-function relationships in, some of these mutants, we also determined the crystal structures of the, CHS C164A, H303Q, and N336A mutants refined to 1.69, 2.0, and 2.15 A, resolution, respectively. The structure of the C164A mutant reveals that, the proposed oxyanion hole formed by His303 and Asn336 remains, undisturbed, allowing this mutant to catalyze malonyl-CoA decarboxylation, without chalcone formation. The structures of the H303Q and N336A mutants, support the importance of His303 and Asn336 in polarizing the thioester, carbonyl of malonyl-CoA during the decarboxylation reaction. In addition, both of these residues may also participate in stabilizing the tetrahedral, transition state during polyketide elongation. Conservation of the, catalytic functions of the active-site residues may occur across a wide, variety of condensing enzymes, including other polyketide and fatty acid, synthases. | + | <StructureSection load='1d6h' size='340' side='right'caption='[[1d6h]], [[Resolution|resolution]] 2.15Å' scene=''> |
| | + | == Structural highlights == |
| | + | <table><tr><td colspan='2'>[[1d6h]] is a 1 chain structure with sequence from [https://en.wikipedia.org/wiki/Medicago_sativa Medicago sativa]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1D6H OCA]. For a <b>guided tour on the structure components</b> use [https://proteopedia.org/fgij/fg.htm?mol=1D6H FirstGlance]. <br> |
| | + | </td></tr><tr id='method'><td class="sblockLbl"><b>[[Empirical_models|Method:]]</b></td><td class="sblockDat" id="methodDat">X-ray diffraction, [[Resolution|Resolution]] 2.15Å</td></tr> |
| | + | <tr id='ligand'><td class="sblockLbl"><b>[[Ligand|Ligands:]]</b></td><td class="sblockDat" id="ligandDat"><scene name='pdbligand=COA:COENZYME+A'>COA</scene>, <scene name='pdbligand=CSD:3-SULFINOALANINE'>CSD</scene>, <scene name='pdbligand=SO4:SULFATE+ION'>SO4</scene></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=1d6h FirstGlance], [http://oca.weizmann.ac.il/oca-bin/ocaids?id=1d6h OCA], [https://pdbe.org/1d6h PDBe], [https://www.rcsb.org/pdb/explore.do?structureId=1d6h RCSB], [https://www.ebi.ac.uk/pdbsum/1d6h PDBsum], [https://prosat.h-its.org/prosat/prosatexe?pdbcode=1d6h ProSAT]</span></td></tr> |
| | + | </table> |
| | + | == Function == |
| | + | [https://www.uniprot.org/uniprot/CHS2_MEDSA CHS2_MEDSA] The primary product of this enzyme is 4,2',4',6'-tetrahydroxychalcone (also termed naringenin-chalcone or chalcone) which can under specific conditions spontaneously isomerize into naringenin.<ref>PMID:10653632</ref> <ref>PMID:11732902</ref> <ref>PMID:11959984</ref> <ref>PMID:15380179</ref> |
| | + | == Evolutionary Conservation == |
| | + | [[Image:Consurf_key_small.gif|200px|right]] |
| | + | Check<jmol> |
| | + | <jmolCheckbox> |
| | + | <scriptWhenChecked>; select protein; define ~consurf_to_do selected; consurf_initial_scene = true; script "/wiki/ConSurf/d6/1d6h_consurf.spt"</scriptWhenChecked> |
| | + | <scriptWhenUnchecked>script /wiki/extensions/Proteopedia/spt/initialview03.spt</scriptWhenUnchecked> |
| | + | <text>to colour the structure by Evolutionary Conservation</text> |
| | + | </jmolCheckbox> |
| | + | </jmol>, as determined by [http://consurfdb.tau.ac.il/ ConSurfDB]. You may read the [[Conservation%2C_Evolutionary|explanation]] of the method and the full data available from [http://bental.tau.ac.il/new_ConSurfDB/main_output.php?pdb_ID=1d6h ConSurf]. |
| | + | <div style="clear:both"></div> |
| | + | <div style="background-color:#fffaf0;"> |
| | + | == Publication Abstract from PubMed == |
| | + | Chalcone synthase (CHS) catalyzes formation of the phenylpropanoid chalcone from one p-coumaroyl-CoA and three malonyl-coenzyme A (CoA) thioesters. The three-dimensional structure of CHS [Ferrer, J.-L., Jez, J. M., Bowman, M. E., Dixon, R. A., and Noel, J. P. (1999) Nat. Struct. Biol. 6, 775-784] suggests that four residues (Cys164, Phe215, His303, and Asn336) participate in the multiple decarboxylation and condensation reactions catalyzed by this enzyme. Here, we functionally characterize 16 point mutants of these residues for chalcone production, malonyl-CoA decarboxylation, and the ability to bind CoA and acetyl-CoA. Our results confirm Cys164's role as the active-site nucleophile in polyketide formation and elucidate the importance of His303 and Asn336 in the malonyl-CoA decarboxylation reaction. We suggest that Phe215 may help orient substrates at the active site during elongation of the polyketide intermediate. To better understand the structure-function relationships in some of these mutants, we also determined the crystal structures of the CHS C164A, H303Q, and N336A mutants refined to 1.69, 2.0, and 2.15 A resolution, respectively. The structure of the C164A mutant reveals that the proposed oxyanion hole formed by His303 and Asn336 remains undisturbed, allowing this mutant to catalyze malonyl-CoA decarboxylation without chalcone formation. The structures of the H303Q and N336A mutants support the importance of His303 and Asn336 in polarizing the thioester carbonyl of malonyl-CoA during the decarboxylation reaction. In addition, both of these residues may also participate in stabilizing the tetrahedral transition state during polyketide elongation. Conservation of the catalytic functions of the active-site residues may occur across a wide variety of condensing enzymes, including other polyketide and fatty acid synthases. |
| | | | |
| - | ==About this Structure==
| + | Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase.,Jez JM, Ferrer JL, Bowman ME, Dixon RA, Noel JP Biochemistry. 2000 Feb 8;39(5):890-902. PMID:10653632<ref>PMID:10653632</ref> |
| - | 1D6H is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Medicago_sativa Medicago sativa] with SO4 and COA as [http://en.wikipedia.org/wiki/ligands ligands]. Active as [http://en.wikipedia.org/wiki/Naringenin-chalcone_synthase Naringenin-chalcone synthase], with EC number [http://www.brenda-enzymes.info/php/result_flat.php4?ecno=2.3.1.74 2.3.1.74] Full crystallographic information is available from [http://ispc.weizmann.ac.il/oca-bin/ocashort?id=1D6H OCA].
| + | |
| | | | |
| - | ==Reference==
| + | From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.<br> |
| - | Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase., Jez JM, Ferrer JL, Bowman ME, Dixon RA, Noel JP, Biochemistry. 2000 Feb 8;39(5):890-902. PMID:[http://ispc.weizmann.ac.il//pmbin/getpm?pmid=10653632 10653632]
| + | </div> |
| - | [[Category: Medicago sativa]]
| + | <div class="pdbe-citations 1d6h" style="background-color:#fffaf0;"></div> |
| - | [[Category: Naringenin-chalcone synthase]]
| + | |
| - | [[Category: Single protein]]
| + | |
| - | [[Category: Bowman, M.E.]]
| + | |
| - | [[Category: Dixon, R.A.]]
| + | |
| - | [[Category: Ferrer, J.L.]]
| + | |
| - | [[Category: Jez, J.M.]]
| + | |
| - | [[Category: Noel, J.P.]]
| + | |
| - | [[Category: COA]]
| + | |
| - | [[Category: SO4]]
| + | |
| - | [[Category: flavonoid biosynthesis]]
| + | |
| - | [[Category: malonyl-coa decarboxylation]]
| + | |
| - | [[Category: polypetide synthase]]
| + | |
| - | [[Category: site-directed mutant]]
| + | |
| | | | |
| - | ''Page seeded by [http://ispc.weizmann.ac.il/oca OCA ] on Tue Nov 20 13:02:31 2007''
| + | ==See Also== |
| | + | *[[Chalcone synthase|Chalcone synthase]] |
| | + | == References == |
| | + | <references/> |
| | + | __TOC__ |
| | + | </StructureSection> |
| | + | [[Category: Large Structures]] |
| | + | [[Category: Medicago sativa]] |
| | + | [[Category: Bowman ME]] |
| | + | [[Category: Dixon RA]] |
| | + | [[Category: Ferrer JL]] |
| | + | [[Category: Jez JM]] |
| | + | [[Category: Noel JP]] |
| Structural highlights
Function
CHS2_MEDSA The primary product of this enzyme is 4,2',4',6'-tetrahydroxychalcone (also termed naringenin-chalcone or chalcone) which can under specific conditions spontaneously isomerize into naringenin.[1] [2] [3] [4]
Evolutionary Conservation
Check, as determined by ConSurfDB. You may read the explanation of the method and the full data available from ConSurf.
Publication Abstract from PubMed
Chalcone synthase (CHS) catalyzes formation of the phenylpropanoid chalcone from one p-coumaroyl-CoA and three malonyl-coenzyme A (CoA) thioesters. The three-dimensional structure of CHS [Ferrer, J.-L., Jez, J. M., Bowman, M. E., Dixon, R. A., and Noel, J. P. (1999) Nat. Struct. Biol. 6, 775-784] suggests that four residues (Cys164, Phe215, His303, and Asn336) participate in the multiple decarboxylation and condensation reactions catalyzed by this enzyme. Here, we functionally characterize 16 point mutants of these residues for chalcone production, malonyl-CoA decarboxylation, and the ability to bind CoA and acetyl-CoA. Our results confirm Cys164's role as the active-site nucleophile in polyketide formation and elucidate the importance of His303 and Asn336 in the malonyl-CoA decarboxylation reaction. We suggest that Phe215 may help orient substrates at the active site during elongation of the polyketide intermediate. To better understand the structure-function relationships in some of these mutants, we also determined the crystal structures of the CHS C164A, H303Q, and N336A mutants refined to 1.69, 2.0, and 2.15 A resolution, respectively. The structure of the C164A mutant reveals that the proposed oxyanion hole formed by His303 and Asn336 remains undisturbed, allowing this mutant to catalyze malonyl-CoA decarboxylation without chalcone formation. The structures of the H303Q and N336A mutants support the importance of His303 and Asn336 in polarizing the thioester carbonyl of malonyl-CoA during the decarboxylation reaction. In addition, both of these residues may also participate in stabilizing the tetrahedral transition state during polyketide elongation. Conservation of the catalytic functions of the active-site residues may occur across a wide variety of condensing enzymes, including other polyketide and fatty acid synthases.
Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase.,Jez JM, Ferrer JL, Bowman ME, Dixon RA, Noel JP Biochemistry. 2000 Feb 8;39(5):890-902. PMID:10653632[5]
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.
See Also
References
- ↑ Jez JM, Ferrer JL, Bowman ME, Dixon RA, Noel JP. Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase. Biochemistry. 2000 Feb 8;39(5):890-902. PMID:10653632
- ↑ Jez JM, Bowman ME, Noel JP. Structure-guided programming of polyketide chain-length determination in chalcone synthase. Biochemistry. 2001 Dec 11;40(49):14829-38. PMID:11732902
- ↑ Jez JM, Bowman ME, Noel JP. Expanding the biosynthetic repertoire of plant type III polyketide synthases by altering starter molecule specificity. Proc Natl Acad Sci U S A. 2002 Apr 16;99(8):5319-24. PMID:11959984 doi:10.1073/pnas.082590499
- ↑ Austin MB, Bowman ME, Ferrer JL, Schroder J, Noel JP. An aldol switch discovered in stilbene synthases mediates cyclization specificity of type III polyketide synthases. Chem Biol. 2004 Sep;11(9):1179-94. PMID:15380179 doi:http://dx.doi.org/10.1016/j.chembiol.2004.05.024
- ↑ Jez JM, Ferrer JL, Bowman ME, Dixon RA, Noel JP. Dissection of malonyl-coenzyme A decarboxylation from polyketide formation in the reaction mechanism of a plant polyketide synthase. Biochemistry. 2000 Feb 8;39(5):890-902. PMID:10653632
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