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• W3169876048 startingPage "5827" @default.
• W3169876048 abstract "Phenylalanine hydroxylase (PAH) deficiency (PAHD) is an autosomal recessive disorder that causes severe injury to the nervous system, the treatment of which mainly depends on dietary therapy. The limited treatment options for PAHD are an incentive to develop new methods to identify more efficient therapeutic drugs, such as agonists which could improve PAH activity. In this study, we aimed to establish a rapid and convenient method for the screening and verification of PAH agonists. We compared fluorospectrophotometry and tandem mass spectrometry for detection of enzymatic formation of tyrosine, finding that the latter was a more sensitive method. We optimized immunoprecipitation purification conditions and measurement conditions of PAH activity. The optimal ratio between PAH protein and magnetic beads was 500 μg protein per 20 μL beads, and the optimized conditions for the detection of PAH enzymatic activity included the presence of 75 μM coenzyme ((6R)-l-erythro-5,6,7,8-tetrahydrobiopterin) and 30 min reaction time. Based on virtual screening, we screened ten candidate agonists from the FDA drug library. Three of these (nefopam, fluocinonide, and risperidone) were found to activate the enzyme in a dose-dependent manner (0.1-10 μM) by the joint method. We tested the efficacy of the three agonists on three PAH mutations (p.I65T, p.H107R, and p.D101N) that influence enzyme activity, and found that risperidone could specifically activate D101N-mutated enzyme. In conclusion, we established a joint method that is highly reliable, cost-effective, labor-saving, and time-saving. And we also found a specific agonist for D101N-mutated PAH by this joint method which may assist the development of clinical treatment for PAHD patients with different enzyme deficiencies." @default.
• W3169876048 created "2021-06-22" @default.
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• W3169876048 date "2021-01-01" @default.
• W3169876048 modified "2023-09-24" @default.
• W3169876048 title "A joint method for the screening of pharmacological chaperones for phenylalanine hydroxylase" @default.
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• W3169876048 doi "https://doi.org/10.1039/d1ob00638j" @default.
• W3169876048 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/34113943" @default.
• W3169876048 hasPublicationYear "2021" @default.
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