LC-MS method for determining the activity of semicarbazide-sensitive amine oxidase in rodents

Wang, Lin; Zhang, Yongqiang; Shuhua, Xiao; Hu, Gaofei; Che, Baoquan; Qing, Hong; Li, Yujuan; Li-xia, Zhuang; Deng, Yulin

doi:10.1039/c2ay05914b

Cited by 6 publications

(2 citation statements)

References 24 publications

(24 reference statements)

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“…Important advantages of LC-MS methods for enzyme activity measurements include a high speed, sensitivity, selectivity, precision, and accuracy. SSAO activity assay based on ESI-MS utilized the oxidation of methylamine as a substrate to formaldehyde (L. Wang et al, 2012). The aldehyde product was subsequently reacted with dopamine to form tetrahydroisoquinoline isomers.…”

Section: Ms-based Activity Assays-what Is Available?mentioning

confidence: 99%

The use of matrix‐assisted laser desorption/ionization mass spectrometry in enzyme activity assays and its position in the context of other available methods

Šebela

2021

Mass Spectrometry Reviews

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Activity assays are indispensable for studying biochemical properties of enzymes. The purposes of measuring activity are wide ranging from a simple detection of the presence of an enzyme to kinetic experiments evaluating the substrate specificity, reaction mechanisms, and susceptibility to inhibitors. Common activity assay methods include spectroscopy, electrochemical sensors, or liquid chromatography coupled with various detection techniques. This review focuses on the use of matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS) as a growing and modern alternative, which offers high speed of analysis, sensitivity, versatility, possibility of automation, and cost‐effectiveness. It may reveal reaction intermediates, side products or measure more enzymes at once. The addition of an internal standard or calculating the ratios of the substrate and product peak intensities and areas overcome the inherent inhomogeneous distribution of analyte and matrix in the sample spot, which otherwise results in a poor reproducibility. Examples of the application of MALDI‐TOF MS for assaying hydrolases (including peptidases and β‐lactamases for antibiotic resistance tests) and other enzymes are provided. Concluding remarks summarize advantages and challenges coming from the present experience, and draw future perspectives such as a screening of large libraries of chemical compounds for their substrate or inhibitory properties towards enzymes.

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Section: Ms-based Activity Assays-what Is Available?mentioning

confidence: 99%

The use of matrix‐assisted laser desorption/ionization mass spectrometry in enzyme activity assays and its position in the context of other available methods

Šebela

2021

Mass Spectrometry Reviews

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“…[20][21][22][23][24][25] Based on different copolymers and chemical bonds, some researchers developed some different polymeric prodrugs focusing on the target, pH-sensitivity, and thermosensitivity. [26][27][28][29][30][31][32] Xiuli Hu and coworkers synthesized a pH-sensitive carrier micelle self-assembled from PEG-PLA. 33 This block copolymer is amphiphilic and can form micelles in an aqueous system with the PLA block in the core and mPEG in the shell (as shown in Scheme 1).…”

Section: Introductionmentioning

confidence: 99%

Studies on the biological character of a new pH-sensitive doxorubicin prodrug with tumor targeting using a LC-MS/MS method

Zhang

Song

et al. 2014

Anal. Methods

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Arterial vascular cell line expressing SSAO: a new tool to study the pathophysiology of vascular amine oxidases

et al. 2013

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Semicarbazide-sensitive amine oxidase (SSAO) widely exists in nature, mainly expressed at significant levels in vasculature. It plays a detrimental role in vascular diseases, particularly atherosclerosis, which occurs mainly in arteries. Herein we for the first time present SSAO expression in arterial lineage of vascular cell line, i.e., human umbilical arterial endothelial cell (HUAEC). Firstly, two commercially available gene transfection reagents were compared to determine high transfection efficiency and then the expression behavior of HUAEC:SSAO was characterized. Furthermore, our model was also been compared with commonly used human embryonic kidney (HEK) cell transfected with the same vector. For enzymatic assay, an in-house developed highly sensitive high performance liquid chromatography electron spray ionization mass spectrometry method was applied. Results indicated that the maximal transfection efficiency in HUAEC was detected by JetPEI™ and transfected protein was expressed at membrane and cytosol of different clones. No significant variations were observed in HUAEC between cell passages 1 and 7, although HEK cell displayed twofold higher SSAO expression level than HUAEC. The transfected SSAO was shown to be released into the cell-culture medium. Both cellular and released types of SSAO exhibited monomer and dimer structural forms. The cytotoxicity determination exhibited large number of viable cells after transfection with JetPEI™. Differential expression characterization of this new cell line demonstrates the correct behavior of SSAO in arterial endothelial cells and also provides a real physiological environment to elucidate the unclear role of this enzyme. In addition, our cellular model could partly solve the problems raised by the loss of enzyme expression found in cultured endothelial cells. This model could also be a useful tool for proteomic base study, screening of interacting protein and analysis of compounds that could modify its activity for therapeutic purposes.

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LC-MS method for determining the activity of semicarbazide-sensitive amine oxidase in rodents

Cited by 6 publications

References 24 publications

The use of matrix‐assisted laser desorption/ionization mass spectrometry in enzyme activity assays and its position in the context of other available methods

The use of matrix‐assisted laser desorption/ionization mass spectrometry in enzyme activity assays and its position in the context of other available methods

Studies on the biological character of a new pH-sensitive doxorubicin prodrug with tumor targeting using a LC-MS/MS method

Arterial vascular cell line expressing SSAO: a new tool to study the pathophysiology of vascular amine oxidases

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