1mdl

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{{STRUCTURE_1mdl| PDB=1mdl | SCENE= }}
{{STRUCTURE_1mdl| PDB=1mdl | SCENE= }}
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'''MANDELATE RACEMASE MUTANT K166R CO-CRYSTALLIZED WITH (R)-MANDELATE'''
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===MANDELATE RACEMASE MUTANT K166R CO-CRYSTALLIZED WITH (R)-MANDELATE===
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==Overview==
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On the basis of the available high-resolution structures of mandelate racemase (MR) from Pseudomonas putida [Landro, J. A., Gerlt, J. A., Kozarich, J. W., Koo, C. W., Shah, V. J., Kenyon, G. L., Neidhart, D. J., Fujita, J., &amp; Petsko, G. A. (1994) Biochemistry 33, 635-643], Lys 166 and His 297 are positioned appropriately to participate in catalysis as acid/base catalysts that either abstract the alpha-proton from the enantiomers of mandelate to form an enolic intermediate or protonate the enolic intermediate to form the enantiomers of mandelate, with Lys 166 participating as the (S)-specific acid/base catalyst and His 297 participating as the (R)-specific acid/base catalyst. In this paper we report the structural and mechanistic properties of the mutant in which Lys 166 has been replaced with arginine (K166R). The structure of K166R has been determined at 1.85 A resolution with the substrate (S)-mandelate bound in the active site. The structure of this complex reveals no geometric alterations in the active site, with the exception that the longer side chain of Arg 166 is necessarily displaced upward from the position occupied by Lys 166 by steric interactions with the bound substrate. In contrast to the H297N mutant of MR [Landro, J. A., Kallarakal, A. T., Ransom, S. C., Gerlt, J. A., Kozarich, J. W., Neidhart, D. J., &amp; Kenyon, G. L. (1991) Biochemistry 30, 9275-9281], the K166R exhibits low levels of racemase activity [kcat is reduced 5 x 10(3)-fold in the (R)- to (S)-direction and 1 x 10(3)-fold in the (S)- to (R)-direction]. The substrate and solvent deuterium isotope effects support a reaction coordinate for the K166R-catalyzed reaction in which the transition state for interconversion of bound (S)-mandelate and the stabilized enolic intermediate is higher in energy that the transition state for interconversion of bound (R)-mandelate and the stabilized enolic intermediate. The solvent deuterium isotope effect when (S)-mandelate is substrate (2.2 +/- 0.3) supports the proposal that the formation of the enolic intermediate involves partial transfer of a solvent-derived proton from Glu 317 to the substrate as the alpha-proton is abstracted [Mitra, B., Kallarakal, A. T., Kozarich, J. W., Gerlt, J. A., Clifton, J. G., Petsko, G. A., &amp; Kenyon, G. L. (1995) Biochemistry 34, 2777-2787].(ABSTRACT TRUNCATED AT 400 WORDS)
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{{ABSTRACT_PUBMED_7893690}}
==About this Structure==
==About this Structure==
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[[Category: Magnesium]]
[[Category: Magnesium]]
[[Category: Mandelate pathway]]
[[Category: Mandelate pathway]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Wed Jul 2 23:44:30 2008''

Revision as of 20:44, 2 July 2008

Image:1mdl.png

Template:STRUCTURE 1mdl

MANDELATE RACEMASE MUTANT K166R CO-CRYSTALLIZED WITH (R)-MANDELATE

Template:ABSTRACT PUBMED 7893690

About this Structure

1MDL is a Single protein structure of sequence from Pseudomonas aeruginosa. Full crystallographic information is available from OCA.

Reference

Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the K166R mutant., Kallarakal AT, Mitra B, Kozarich JW, Gerlt JA, Clifton JG, Petsko GA, Kenyon GL, Biochemistry. 1995 Mar 7;34(9):2788-97. PMID:7893690

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