Isotope Probing of the UDP-Apiose/UDP-Xylose Synthase Reaction: Evidence of a Mechanism via a Coupled Oxidation and Aldol Cleavage
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- 作者：Dr. Thomas Eixelsberger ; Doroteja Horvat ; Dr. Alexander Gutmann ; Prof. Dr. Hansjö ; rg Weber and Prof. Dr. Bernd Nidetzky
- 刊名：Angewandte Chemie International Edition
- 出版年：2017
- 出版时间：February 20, 2017
- 年：2017
- 卷：56
- 期：9
- 页码：2503-2507
- 全文大小：1614K
- ISSN：1521-3773
文摘
The C-branched sugar d-apiose (Api) is essential for plant cell-wall development. An enzyme-catalyzed decarboxylation/pyranoside ring-contraction reaction leads from UDP-α-d-glucuronic acid (UDP-GlcA) to the Api precursor UDP-α-d-apiose (UDP-Api). We examined the mechanism of UDP-Api/UDP-α-d-xylose synthase (UAXS) with site-selectively 2H-labeled and deoxygenated substrates. The analogue UDP-2-deoxy-GlcA, which prevents C-2/C-3 aldol cleavage as the plausible initiating step of pyranoside-to-furanoside conversion, did not give the corresponding Api product. Kinetic isotope effects (KIEs) support an UAXS mechanism in which substrate oxidation by enzyme-NAD+ and retro-aldol sugar ring-opening occur coupled in a single rate-limiting step leading to decarboxylation. Rearrangement and ring-contracting aldol addition in an open-chain intermediate then give the UDP-Api aldehyde, which is intercepted via reduction by enzyme-NADH.