Enzymatic Synthesis of Isotopically Labeled Hydrogen Peroxide for Mass Spectrometry-Based Applications.

IF 3.1 2区 化学 Q2 BIOCHEMICAL RESEARCH METHODS Journal of the American Society for Mass Spectrometry Pub Date : 2024-12-04 Epub Date: 2024-10-04 DOI:10.1021/jasms.4c00326
Margaret Hoare, Ruiyue Tan, Isabella Militi, Kevin A Welle, Kyle Swovick, Jennifer R Hryhorenko, Sina Ghaemmaghami
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Abstract

Methionine oxidation is involved in multiple biological processes including protein misfolding and enzyme regulation. However, it is often challenging to measure levels of methionine oxidation by mass spectrometry, in part due to the prevalence of artifactual oxidation that occurs during the sample preparation and ionization steps of typical proteomic workflows. Isotopically labeled hydrogen peroxide (H218O2) can be used to block unoxidized methionines and enables accurate measurement of in vivo levels of methionine oxidation. However, H218O2 is an expensive reagent that can be difficult to obtain from commercial sources. Here, we report a method for synthesizing H218O2 in-house. Glucose oxidase catalyzes the oxidation of β-d-glucose and produces hydrogen peroxide in the process. We took advantage of this reaction to enzymatically synthesize H218O2 from 18O2 and assessed its concentration, purity, and utility in measuring methionine oxidation levels by mass spectrometry.

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基于质谱应用的同位素标记过氧化氢的酶法合成。
蛋氨酸氧化参与多种生物过程,包括蛋白质错误折叠和酶调控。然而,用质谱法测量蛋氨酸氧化水平往往具有挑战性,部分原因是在典型蛋白质组学工作流程的样品制备和电离步骤中普遍存在人为氧化现象。同位素标记的过氧化氢(H218O2)可用于阻断未氧化的蛋氨酸,从而准确测量体内蛋氨酸的氧化水平。然而,H218O2 是一种昂贵的试剂,很难从商业渠道获得。在此,我们报告了一种内部合成 H218O2 的方法。葡萄糖氧化酶催化β-d-葡萄糖氧化,并在此过程中产生过氧化氢。我们利用这一反应从 18O2 酶法合成了 H218O2,并评估了其浓度、纯度以及通过质谱法测量蛋氨酸氧化水平的实用性。
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来源期刊
CiteScore
5.50
自引率
9.40%
发文量
257
审稿时长
1 months
期刊介绍: The Journal of the American Society for Mass Spectrometry presents research papers covering all aspects of mass spectrometry, incorporating coverage of fields of scientific inquiry in which mass spectrometry can play a role. Comprehensive in scope, the journal publishes papers on both fundamentals and applications of mass spectrometry. Fundamental subjects include instrumentation principles, design, and demonstration, structures and chemical properties of gas-phase ions, studies of thermodynamic properties, ion spectroscopy, chemical kinetics, mechanisms of ionization, theories of ion fragmentation, cluster ions, and potential energy surfaces. In addition to full papers, the journal offers Communications, Application Notes, and Accounts and Perspectives
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