1i2n: Difference between revisions

Revision as of 19:29, 2 May 2008

CRYSTAL STRUCTURE OF ESCHERICHIA COLI TRANSALDOLASE B MUTANT N35A

OverviewOverview

The roles of invariant residues at the active site of transaldolase B from Escherichia coli have been probed by site-directed mutagenesis. The mutant enzymes D17A, N35A, E96A, T156A, and S176A were purified from a talB-deficient host and analyzed with respect to their 3D structure and kinetic behavior. X-ray analysis showed that side chain replacement did not induce unanticipated structural changes in the mutant enzymes. Three mutations, N35A, E96A, and T156A resulted mainly in an effect on apparent kcat, with little changes in apparent Km values for the substrates. Residues N35 and T156 are involved in the positioning of a catalytic water molecule at the active site and the side chain of E96 participates in concert with this water molecule in proton transfer during catalysis. Substitution of Ser176 by alanine resulted in a mutant enzyme with 2.5% residual activity. The apparent Km value for the donor substrate, fructose 6-phosphate, was increased nearly fivefold while the apparent Km value for the acceptor substrate, erythrose 4-phosphate remained unchanged, consistent with a function for S176 in the binding of the C1 hydroxyl group of the donor substrate. The mutant D17A showed a 300-fold decrease in kcat, and a fivefold increase in the apparent Km value for the acceptor substrate erythrose 4-phosphate, suggesting a role of this residue in carbon-carbon bond cleavage and stabilization of the carbanion/enamine intermediate.

About this StructureAbout this Structure

1I2N is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.

ReferenceReference

Identification of catalytically important residues in the active site of Escherichia coli transaldolase., Schorken U, Thorell S, Schurmann M, Jia J, Sprenger GA, Schneider G, Eur J Biochem. 2001 Apr;268(8):2408-15. PMID:11298760 Page seeded by OCA on Fri May 2 19:29:55 2008

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OCA

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 [[Image:1i2n.jpg|left|200px]]
- {{Structure
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 '''CRYSTAL STRUCTURE OF ESCHERICHIA COLI TRANSALDOLASE B MUTANT N35A'''
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 [[Category: Schneider, G.]]
 [[Category: Thorell, S.]]
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+''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May  2 19:29:55 2008''
-''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun Mar 30 21:13:43 2008''