3P74

H181N mutant of pentaerythritol tetranitrate reductase containing a C-terminal His8-tag

Summary for 3P74

• Entry DOI: 10.2210/pdb3p74/pdb
• Related: 1H50
• Descriptor: Pentaerythritol tetranitrate reductase, FLAVIN MONONUCLEOTIDE (3 entities in total)
• Functional Keywords: old yellow enzyme family, alpha/beta barrel, oxidoreductase
• Biological source: Enterobacter cloacae
• Total number of polymer chains: 1
• Total formula weight: 41072.66

Authors

Toogood, H.S.,Scrutton, N.S. (deposition date: 2010-10-12, release date: 2011-03-23, Last modification date: 2024-02-21)

Primary citation

Toogood, H.S.,Fryszkowska, A.,Hulley, M.,Sakuma, M.,Mansell, D.,Stephens, G.M.,Gardiner, J.M.,Scrutton, N.S.
A Site-Saturated Mutagenesis Study of Pentaerythritol Tetranitrate Reductase Reveals that Residues 181 and 184 Influence Ligand Binding, Stereochemistry and Reactivity.
Chembiochem, 12:738-749, 2011

PubMed Abstract: We have conducted a site-specific saturation mutagenesis study of H181 and H184 of flavoprotein pentaerythritol tetranitrate reductase (PETN reductase) to probe the role of these residues in substrate binding and catalysis with a variety of α,β-unsaturated alkenes. Single mutations at these residues were sufficient to dramatically increase the enantiopurity of products formed by reduction of 2-phenyl-1-nitropropene. In addition, many mutants exhibited a switch in reactivity to predominantly catalyse nitro reduction, as opposed to CC reduction. These mutants showed an enhancement in a minor side reaction and formed 2-phenylpropanal oxime from 2-phenyl-1-nitropropene. The multiple binding conformations of hydroxy substituted nitro-olefins in PETN reductase were examined by using both structural and catalytic techniques. These compounds were found to bind in both active and inhibitory complexes; this highlights the plasticity of the active site and the ability of the H181/H184 couple to coordinate with multiple functional groups. These properties demonstrate the potential to use PETN reductase as a scaffold in the development of industrially useful biocatalysts.

PubMed: 21374779
DOI: 10.1002/cbic.201000662

Experimental method

X-RAY DIFFRACTION (1.2 Å)