Mechanism-based inhibition of an aldolase at high concentrations of its natural substrate acetaldehyde: structural insights and protective strategies

Chemical Science
Markus DickJörg Pietruszka

Abstract

2-Deoxy-d-ribose-5-phosphate aldolase (DERA) is used in organic synthesis for the enantioselective reaction between acetaldehyde and a broad range of other aldehydes as acceptor molecules. Nevertheless, its application is hampered by a poor tolerance towards high concentrations of acetaldehyde, its natural substrate. While numerous studies have been performed searching for new, more acetaldehyde-resistant DERAs, the mechanism underlying this deactivation process has remained elusive. By using NMR spectroscopy on both the protein and the small-molecule scale, we could show that a reaction product binds to the inner part of the enzyme, and that this effect can be partly reversed via heating. The crystal structure of DERA before and after acetaldehyde incubation was determined at high resolution, revealing a covalently bound reaction product bridging the catalytically active lysine (K167) to a nearby cysteine (C47) in the deactivated enzyme. A reaction mechanism is proposed where crotonaldehyde as the aldol product of two acetaldehyde molecules after water elimination forms a Schiff base with the lysine side chain, followed by Michael addition of the cysteine thiol group to the Cβ atom of the inhibitor. In support of this mechanis...Continue Reading

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Citations

Dec 7, 2019·The Journal of Biological Chemistry·Taeho KimAlexander F Yakunin
Oct 5, 2018·Applied Microbiology and Biotechnology·Meera HaridasUlf Hanefeld
Mar 24, 2021·Proceedings of the National Academy of Sciences of the United States of America·Andrew D HansonA Harvey Millar
Aug 20, 2021·Applied Microbiology and Biotechnology·Juha RouvinenAnu Koivula
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Dec 21, 2021·Chemical Society Reviews·Ulf HanefeldCaroline E Paul
Jan 21, 2022·Chembiochem : a European Journal of Chemical Biology·Andreas KunzendorfGerrit J Poelarends

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Methods Mentioned

BETA
NMR
X-ray
size exclusion chromatography

Software Mentioned

DSSP
NMRPipe
POVScript
Coot
SPARTA
CSI
MolProbity
Raster3D
phenix
MOLREP

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