1eb8

Categories: Manihot esculenta | Single protein | Transferred entry: 3.3.2.4 | Effenberger, F. | Foerster, S. | Kobler, C. | Lauble, H. | Miehlich, B. | Wajant, H. | MPD | Active-site tunnel mutant | Hydroxynitrile lyase | Lyase | Substrate specificity

STRUCTURE DETERMINANTS OF SUBSTRATE SPECIFICITY OF HYDROXYNITRILE LYASE FROM MANIHOT ESCULENTA

Overview

Tryptophan 128 of hydroxynitrile lyase of Manihot esculenta (MeHNL) covers, a significant part of a hydrophobic channel that gives access to the, active site of the enzyme. This residue was therefore substituted in the, mutant MeHNL-W128A by alanine to study its importance for the substrate, specificity of the enzyme. Wild-type MeHNL and MeHNL-W128A showed, comparable activity on the natural substrate acetone cyanohydrin (53 and, 40 U/mg, respectively). However, the specific activities of MeHNL-W128A, for the unnatural substrates mandelonitrile and 4-hydroxymandelonitrile, are increased 9-fold and approximately 450-fold, respectively, compared, with the wild-type MeHNL. The crystal structure of the MeHNL-W128A, substrate-free form at 2.1 A resolution indicates that the W128A, substitution has significantly enlarged the active-site channel entrance, and thereby explains the observed changes in substrate specificity for, bulky substrates. Surprisingly, the MeHNL-W128A--4-hydroxybenzaldehyde, complex structure at 2.1 A resolution shows the presence of two, hydroxybenzaldehyde molecules in a sandwich type arrangement in the active, site with an additional hydrogen bridge to the reacting center.

About this Structure

1EB8 is a Single protein structure of sequence from Manihot esculenta with MPD as ligand. Active as Transferred entry: 3.3.2.4, with EC number 4.2.1.37 Known structural/functional Site: . Full crystallographic information is available from OCA.

Reference

Structure determinants of substrate specificity of hydroxynitrile lyase from Manihot esculenta., Lauble H, Miehlich B, Forster S, Kobler C, Wajant H, Effenberger F, Protein Sci. 2002 Jan;11(1):65-71. PMID:11742123

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