Crystallographic and spectroscopic snapshots reveal a dehydrogenase in action.

Lu Huo,Babak Andi,Allen M Orville,Shingo Esaki,Hiroaki Iwaki,Ian Davis,Fange Liu,Yoshie Hasegawa,Aimin Liu

doi:10.1038/ncomms6935

Abstract

Aldehydes are ubiquitous intermediates in metabolic pathways and their innate reactivity can often make them quite unstable. There are several aldehydic intermediates in the metabolic pathway for tryptophan degradation that can decay into neuroactive compounds that have been associated with numerous neurological diseases. An enzyme of this pathway, 2-aminomuconate-6-semialdehyde dehydrogenase, is responsible for ‘disarming’ the final aldehydic intermediate. Here we show the crystal structures of a bacterial analogue enzyme in five catalytically relevant forms: resting state, one binary and two ternary complexes, and a covalent, thioacyl intermediate. We also report the crystal structures of a tetrahedral, thiohemiacetal intermediate, a thioacyl intermediate and an NAD+-bound complex from an active site mutant. These covalent intermediates are characterized by single-crystal and solution-state electronic absorption spectroscopy. The crystal structures reveal that the substrate undergoes an E/Z isomerization at the enzyme active site before an sp3-to-sp2 transition during enzyme-mediated oxidation.

Highlights

Aldehydes are ubiquitous intermediates in metabolic pathways and their innate reactivity can often make them quite unstable
The structure of aminomuconate-6-semialdehyde dehydrogenase (AMSDH) will help to address questions such as what contributes to substrate specificity for the semialdehyde dehydrogenase and how 2-AMS is bound and activated during catalysis
In a coupled-enzyme assay, amino b-carboxymuconate e-semialdehyde decarboxylase (ACMSD), AMSDH and NAD þ are included in the reaction system

Summary

Introduction

Aldehydes are ubiquitous intermediates in metabolic pathways and their innate reactivity can often make them quite unstable. There are several aldehydic intermediates in the metabolic pathway for tryptophan degradation that can decay into neuroactive compounds that have been associated with numerous neurological diseases. An enzyme of this pathway, 2-aminomuconate-6-semialdehyde dehydrogenase, is responsible for ‘disarming’ the final aldehydic intermediate. The concentration of quinolinic acid, a nonenzymatically derived decay product of an intermediate of the kynurenine pathway used for NAD þ biosynthesis, is elevated over 20-fold in patients’ cerebrospinal fluid with AIDS dementia complex, aseptic meningitis, opportunistic infections or neoplasms[7], and more than 300-fold in the brain of human immunodeficiency virus-infected patients[8].

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Conclusion