A d-2-hydroxyglutarate biosensor based on specific transcriptional regulator DhdR
Dan Xiao, Wen Zhang, Xiaoting Guo, Yidong Liu, Chunxia Hu, Shiting Guo, Zhaoqi Kang, Xianzhi Xu, Cuiqing Ma, Chao Gao, Ping Xu
Abstract
D-2-Hydroxyglutarate (D-2-HG) is a metabolite involved in many physiological metabolic processes. When D-2-HG is aberrantly accumulated due to mutations in isocitrate dehydrogenase or D-2-HG dehydrogenase, it functions in a pro-oncogenic manner and is thus considered a therapeutic target and biomarker in many cancers. In this study, DhdR from Achromobacter denitrificans NBRC 15125 is identified as an allosteric transcriptional factor that negatively regulates D-2-HG dehydrogenase expression and responds to the presence of D-2-HG. Based on the allosteric effect of DhdR, a D-2-HG biosensor is developed by combining DhdR with amplified luminescent proximity homogeneous assay (AlphaScreen) technology. The biosensor is able to detect D-2-HG in serum, urine, and cell culture medium with high specificity and sensitivity. Additionally, this biosensor is used to identify the role of D-2-HG metabolism in lipopolysaccharide biosynthesis of Pseudomonas aeruginosa, demonstrating its broad usages.