2fhn

From Proteopedia

Revision as of 14:52, 20 March 2008

Avidin related protein AVR4 (C122S, K109I mutant) in complex with BNA

Overview

The hydrolysis of biotinyl p-nitrophenyl ester (BNP) by a series of avidin derivatives was examined. Surprisingly, a hyperthermostable avidin-related protein (AVR4) was shown to display extraordinary yet puzzling hydrolytic activity. In order to evaluate the molecular determinants that contribute to the reaction, the crystal structure of AVR4 was compared with those of avidin, streptavidin and key mutants of the two proteins in complex with biotinyl p-nitroanilide (BNA), the inert amide analogue of BNP. The structures revealed that a critical lysine residue contributes to the hydrolysis of BNP by avidin but has only a minor contribution to the AVR4-mediated reaction. Indeed, the respective rates of hydrolysis among the different avidins reflect several molecular parameters, including binding-site architecture, the availability of the ligand to solvent and the conformation of the ligand and consequent susceptibility to efficient nucleophilic attack. In avidin, the interaction of BNP with Lys111 and disorder of the L3,4 loop (and consequent solvent availability) together comprise the major driving force behind the hydrolysis, whereas in AVR4 the status of the ligand (the pseudo-substrate) is a major distinguishing feature. In the latter protein, a unique conformation of the L3,4 loop restrains the pseudo-substrate, thereby exposing the carbonyl carbon atom to nucleophilic attack. In addition, due to its conformation, the pseudo-substrate in the AVR4 complex cannot interact with the conserved lysine analogue (Lys109); instead, this function is superseded by polar interactions with Arg112. The results demonstrate that, in highly similar proteins, different residues can perform the same function and that subtle differences in the active-site architecture of such proteins can result in alternative modes of reaction.

About this Structure

2FHN is a Single protein structure of sequence from Gallus gallus. Full crystallographic information is available from OCA.

Reference

Factors dictating the pseudocatalytic efficiency of avidins., Prizant M, Eisenberg-Domovich Y, Hytonen VP, Kulomaa MS, Wilchek M, Bayer EA, Livnah O, J Mol Biol. 2006 May 5;358(3):754-63. Epub 2006 Mar 3. PMID:16546211

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