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| {{STRUCTURE_1fnk| PDB=1fnk | SCENE= }} | | {{STRUCTURE_1fnk| PDB=1fnk | SCENE= }} |
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| '''CRYSTAL STRUCTURE ANALYSIS OF CHORISMATE MUTASE MUTANT C88K/R90S'''
| | ===CRYSTAL STRUCTURE ANALYSIS OF CHORISMATE MUTASE MUTANT C88K/R90S=== |
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| ==Overview==
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| Combinatorial mutagenesis and in vivo selection experiments previously afforded functional variants of the AroH class Bacillus subtilis chorismate mutase lacking the otherwise highly conserved active site residue Arg(90). Here, we present a detailed kinetic and crystallographic study of several such variants. Removing the arginine side chain (R90G and R90A) reduced catalytic efficiency by more than 5 orders of magnitude. Reintroducing a positive charge to the active site through lysine substitutions restored more than a factor of a thousand in k(cat). Remarkably, the lysine could be placed at position 90 or at the more remote position 88 provided a sterically suitable residue was present at the partner site. Crystal structures of the double mutants C88S/R90K and C88K/R90S show that the lysine adopts an extended conformation that would place its epsilon-ammonium group within hydrogen-bonding distance of the ether oxygen of bound chorismate in the transition state. These results provide support for the hypothesis that developing negative charge in the highly polarized transition state is stabilized electrostatically by a strategically placed cation. The implications of this finding for the mechanism of all natural chorismate mutases and for the design of artificial catalysts are discussed.
| | The line below this paragraph, {{ABSTRACT_PUBMED_10960481}}, adds the Publication Abstract to the page |
| | (as it appears on PubMed at http://www.pubmed.gov), where 10960481 is the PubMed ID number. |
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| | {{ABSTRACT_PUBMED_10960481}} |
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| ==About this Structure== | | ==About this Structure== |
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| [[Category: Pseudo-alpha beta-barrel]] | | [[Category: Pseudo-alpha beta-barrel]] |
| [[Category: Trimer]] | | [[Category: Trimer]] |
| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 16:32:26 2008'' | | |
| | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Tue Jul 1 03:37:27 2008'' |