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| | {{STRUCTURE_2dw6| PDB=2dw6 | SCENE= }} | | {{STRUCTURE_2dw6| PDB=2dw6 | SCENE= }} |
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| - | '''Crystal structure of the mutant K184A of D-Tartrate Dehydratase from Bradyrhizobium japonicum complexed with Mg++ and D-tartrate'''
| + | ===Crystal structure of the mutant K184A of D-Tartrate Dehydratase from Bradyrhizobium japonicum complexed with Mg++ and D-tartrate=== |
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| - | ==Overview==
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| - | We focus on the assignment of function to and elucidation of structure-function relationships for a member of the mechanistically diverse enolase superfamily encoded by the Bradyrhizobium japonicum genome (bll6730; GI:27381841). As suggested by sequence alignments, the active site contains the same functional groups found in the active site of mandelate racemase (MR) that catalyzes a 1,1-proton transfer reaction: two acid/base catalysts, Lys 184 at the end of the second beta-strand, and a His 322-Asp 292 dyad at the ends of the seventh and sixth beta-strands, respectively, as well as ligands for an essential Mg2+, Asp 213, Glu 239, and Glu 265 at the ends of the third, fourth, and fifth beta-strands, respectively. We screened a library of 46 acid sugars and discovered that only d-tartrate is dehydrated, yielding oxaloacetate as product. The kinetic constants (kcat = 7.3 s(-1); kcat/KM = 8.5 x 10(4) M(-1) s(-1)) are consistent with assignment of the d-tartrate dehydratase (TarD) function. The kinetic phenotypes of mutants as well as the structures of liganded complexes are consistent with a mechanism in which Lys 184 initiates the reaction by abstraction of the alpha-proton to generate a Mg2+-stabilized enediolate intermediate, and the vinylogous beta-elimination of the 3-OH group is general acid-catalyzed by the His 322, accomplishing the anti-elimination of water. The replacement of the leaving group by solvent-derived hydrogen is stereorandom, suggesting that the enol tautomer of oxaloacetate is the product; this expectation was confirmed by its observation by 1H NMR spectroscopy. Thus, the TarD-catalyzed reaction is a "simple" extension of the two-step reaction catalyzed by MR: base-catalyzed proton abstraction to generate a Mg2+-stabilized enediolate intermediate followed by acid-catalyzed decomposition of that intermediate to yield the product.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_17144653}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 17144653 is the PubMed ID number. |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Enolase superfamily]] | | [[Category: Enolase superfamily]] |
| | [[Category: L-tartrate]] | | [[Category: L-tartrate]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun May 4 01:30:17 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Jul 28 02:34:36 2008'' |
Revision as of 23:34, 27 July 2008
Template:STRUCTURE 2dw6
Crystal structure of the mutant K184A of D-Tartrate Dehydratase from Bradyrhizobium japonicum complexed with Mg++ and D-tartrate
Template:ABSTRACT PUBMED 17144653
About this Structure
2DW6 is a Single protein structure of sequence from Bradyrhizobium japonicum. Full crystallographic information is available from OCA.
Reference
Evolution of enzymatic activities in the enolase superfamily: D-tartrate dehydratase from Bradyrhizobium japonicum., Yew WS, Fedorov AA, Fedorov EV, Wood BM, Almo SC, Gerlt JA, Biochemistry. 2006 Dec 12;45(49):14598-608. PMID:17144653
Page seeded by OCA on Mon Jul 28 02:34:36 2008
