An enzymic l-2-hydroxyglutarate biosensor based on l-2-hydroxyglutarate dehydrogenase from Azoarcus olearius.

l-2-Hydroxyglutarate (l-2-HG) is a critical signaling and immune metabolite but its excessive accumulation can lead to l-2-hydroxyglutaric aciduria, renal cancer, and other diseases. Development of efficient and high-throughput methods for selective l-2-HG detection is urgently required. In this study, l-2-HG dehydrogenase in Azoarcus olearius BH72 (AoL2HGDH) was screened from ten homologs and identified as an enzyme with high specificity and activity toward l-2-HG dehydrogenation. Then, an enzymatic assay-based l-2-HG-sensing fluorescent reporter, EaLHGFR which consists of AoL2HGDH and resazurin, was developed for the detection of l-2-HG. The response magnitude and limit of detection of EaLHGFR were systematically optimized using a single-factor screening strategy. The optimal biosensor EaLHGFR-2 exhibited a response magnitude of 2189.25 ± 26.89% and a limit of detection of 0.042 μM. It can accurately detect the concentration of l-2-HG in bacterial and cellular samples as well as human body fluids. Considering its desirable properties, EaLHGFR-2 may be a promising alternative for quantitation of l-2-HG in biological samples.

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Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.