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1qit

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{{STRUCTURE_1qit| PDB=1qit | SCENE= }}
{{STRUCTURE_1qit| PDB=1qit | SCENE= }}
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'''ASPARTATE AMINOTRANSFERASE FROM ESCHERICHIA COLI, C191W MUTATION, WITH BOUND MALEATE'''
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===ASPARTATE AMINOTRANSFERASE FROM ESCHERICHIA COLI, C191W MUTATION, WITH BOUND MALEATE===
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==Overview==
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In previous kinetic studies of Escherichia coli aspartate aminotransferase, it was determined that some substitutions of conserved cysteine 191, which is located outside of the active site, altered the kinetic parameters of the enzyme (Gloss,L.M., Spencer,D. E. and Kirsch,J.F., 1996, Protein Struct. Funct. Genet., 24, 195-208). The mutations resulted in an alkaline shift of 0.6-0.8 pH units for the pK(a) of the internal aldimine between the PLP cofactor and Lys258. The change in the pK(a) affected the pH dependence of the k(cat)/K(m) (aspartate) values for the mutant enzymes. To help to understand these observations, crystal structures of five mutant forms of E.coli aspartate aminotransferase (the maleate complexes of C191S, C191F, C191Y and C191W, and C191S without maleate) were determined at about 2 A resolution in the presence of the pyridoxal phosphate cofactor. The overall three-dimensional fold of each mutant enzyme is the same as that of the wild-type protein, but there is a rotation of the mutated side chain around its C(alpha)-C(beta) bond. This side chain rotation results in a change in the pattern of hydrogen bonding connecting the mutant residue and the protonated Schiff base of the cofactor, which could account for the altered pK(a) of the Schiff base imine nitrogen that was reported previously. These results demonstrate how residues outside the active site can be important in helping determine the subtleties of the active site amino acid geometries and interactions and how mutations outside the active site can have effects on catalysis. In addition, these results help explain the surprising result previously reported that, for some mutant proteins, replacement of a buried cysteine with an aromatic side chain did not destabilize the protein fold. Instead, rotation around the C(alpha)-C(beta) bond allowed each large aromatic side chain to become buried in a nearby pocket without large changes in the enzyme's backbone geometry.
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{{ABSTRACT_PUBMED_10708649}}
==About this Structure==
==About this Structure==
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[[Category: Maleate]]
[[Category: Maleate]]
[[Category: Pyridoxal phosphate]]
[[Category: Pyridoxal phosphate]]
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sat May 3 06:19:31 2008''
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''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Jul 27 19:00:57 2008''

Revision as of 16:01, 27 July 2008

Image:1qit.png

Template:STRUCTURE 1qit

ASPARTATE AMINOTRANSFERASE FROM ESCHERICHIA COLI, C191W MUTATION, WITH BOUND MALEATE

Template:ABSTRACT PUBMED 10708649

About this Structure

1QIT is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.

Reference

The role of residues outside the active site: structural basis for function of C191 mutants of Escherichia coli aspartate aminotransferase., Jeffery CJ, Gloss LM, Petsko GA, Ringe D, Protein Eng. 2000 Feb;13(2):105-12. PMID:10708649

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