2dw6
From Proteopedia
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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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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:[http://www.ncbi.nlm.nih.gov/pubmed/17144653 17144653] | 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:[http://www.ncbi.nlm.nih.gov/pubmed/17144653 17144653] | ||
[[Category: Bradyrhizobium japonicum]] | [[Category: Bradyrhizobium japonicum]] | ||
| - | [[Category: D(-)-tartrate dehydratase]] | ||
[[Category: Single protein]] | [[Category: Single protein]] | ||
[[Category: Almo, S C.]] | [[Category: Almo, S C.]] | ||
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[[Category: Wood, B M.]] | [[Category: Wood, B M.]] | ||
[[Category: Yew, W S.]] | [[Category: Yew, W S.]] | ||
| - | [[Category: | + | [[Category: D-tartrate]] |
| - | [[Category: | + | [[Category: D-tartrate dehydratase]] |
| - | [[Category: | + | [[Category: Enolase superfamily]] |
| - | [[Category: | + | [[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 | + | |
Revision as of 22:30, 3 May 2008
Crystal structure of the mutant K184A of D-Tartrate Dehydratase from Bradyrhizobium japonicum complexed with Mg++ and D-tartrate
Overview
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.
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 Sun May 4 01:30:17 2008
