Spectroscopic method for measuring activity of cis-aconitate decarboxylase, an important metabolic regulator of immune responses

IF 2.6 4区生物学 Q2 BIOCHEMICAL RESEARCH METHODS

Analytical biochemistry Pub Date : 2025-07-11 DOI:10.1016/j.ab.2025.115944

Kevin Knowlan, Cody L. Hoop, Nadya I. Tarasova

引用次数: 0

Abstract

Cis-aconitate decarboxylase (ACOD1) is a key enzyme converting cis-aconitate to itaconate, which has therapeutic potential for inflammatory diseases. Existing methods to measure ACOD1 activity and itaconate are often expensive and complex. We developed a novel, high-throughput spectrophotometric assay using the Fürth-Herrmann reaction. Our method quantifies ACOD1-catalyzed itaconate production by leveraging distinct absorbance ratios of cis-aconitate and itaconate at 386 nm and 440 nm. We optimized parameters, characterized human ACOD1 kinetics, and determined an IC₅₀ for citraconate consistent with previous reports. This simple, fast, and reliable assay, requiring only a UV–Vis spectrophotometer, will accelerate screening for ACOD1 modulators, speeding up therapeutic development.

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测定免疫反应重要代谢调节因子顺式乌头酸脱羧酶活性的光谱方法

顺式乌头酸脱羧酶（ACOD1）是将顺式乌头酸转化为衣康酸的关键酶，具有治疗炎性疾病的潜力。现有的测量ACOD1活性和衣康酸的方法通常既昂贵又复杂。我们利用 rth- herrmann反应开发了一种新的高通量分光光度法。我们的方法量化acod1催化衣康酸的生产，利用不同的吸光度比的顺- aconate和衣康酸在386 nm和440 nm。我们优化了参数，表征了人类ACOD1动力学，并确定了柠檬酸盐的IC50，与先前的报道一致。这种简单、快速、可靠的检测方法，只需要一个紫外可见分光光度计，将加速ACOD1调节剂的筛选，加快治疗开发。

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来源期刊

Analytical biochemistry 生物-分析化学

CiteScore

5.70

自引率

0.00%

发文量

283

审稿时长

44 days

期刊介绍： The journal''s title Analytical Biochemistry: Methods in the Biological Sciences declares its broad scope: methods for the basic biological sciences that include biochemistry, molecular genetics, cell biology, proteomics, immunology, bioinformatics and wherever the frontiers of research take the field. The emphasis is on methods from the strictly analytical to the more preparative that would include novel approaches to protein purification as well as improvements in cell and organ culture. The actual techniques are equally inclusive ranging from aptamers to zymology. The journal has been particularly active in: -Analytical techniques for biological molecules- Aptamer selection and utilization- Biosensors- Chromatography- Cloning, sequencing and mutagenesis- Electrochemical methods- Electrophoresis- Enzyme characterization methods- Immunological approaches- Mass spectrometry of proteins and nucleic acids- Metabolomics- Nano level techniques- Optical spectroscopy in all its forms. The journal is reluctant to include most drug and strictly clinical studies as there are more suitable publication platforms for these types of papers.