3NNE

Crystal structure of choline oxidase S101A mutant

Summary for 3NNE

• Entry DOI: 10.2210/pdb3nne/pdb
• Related: 2JBV 3LJP
• Descriptor: Choline oxidase, FLAVIN-ADENINE DINUCLEOTIDE, ACETATE ION, ... (4 entities in total)
• Functional Keywords: oxidase, flavoprotein, kinetics, reductive half-reaction, choline, oxidoreductase
• Biological source: Arthrobacter globiformis
• Total number of polymer chains: 8
• Total formula weight: 485818.78

Authors

Wang, Y.-F.,Finnegan, S.,Yuan, H.,Orville, A.M.,Weber, I.T.,Gadda, G. (deposition date: 2010-06-23, release date: 2010-07-28, Last modification date: 2024-10-16)

Primary citation

Finnegan, S.,Yuan, H.,Wang, Y.F.,Orville, A.M.,Weber, I.T.,Gadda, G.
Structural and kinetic studies on the Ser101Ala variant of choline oxidase: Catalysis by compromise.
Arch.Biochem.Biophys., 501:207-213, 2010

PubMed Abstract: The oxidation of choline catalyzed by choline oxidase includes two reductive half-reactions where FAD is reduced by the alcohol substrate and by an aldehyde intermediate transiently formed in the reaction. Each reductive half-reaction is followed by an oxidative half-reaction where the reduced flavin is oxidized by oxygen. Here, we have used mutagenesis to prepare the Ser101Ala mutant of choline oxidase and have investigated the impact of this mutation on the structural and kinetic properties of the enzyme. The crystallographic structure of the Ser101Ala enzyme indicates that the only differences between the mutant and wild-type enzymes are the lack of a hydroxyl group on residue 101 and a more planar configuration of the flavin in the mutant enzyme. Kinetics established that replacement of Ser101 with alanine yields a mutant enzyme with increased efficiencies in the oxidative half-reactions and decreased efficiencies in the reductive half-reactions. This is accompanied by a significant decrease in the overall rate of turnover with choline. Thus, this mutation has revealed the importance of a specific residue for the optimization of the overall turnover of choline oxidase, which requires fine-tuning of four consecutive half-reactions for the conversion of an alcohol to a carboxylic acid.

PubMed: 20561507
DOI: 10.1016/j.abb.2010.06.014

Experimental method

X-RAY DIFFRACTION (2.47 Å)