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Open AccessArticle

Structural and Functional Impact of Seven Missense Variants of Phenylalanine Hydroxylase

1
Department of Molecular Biology, Faculty of Natural Sciences, Comenius University, Ilkovicova 6, 842 15 Bratislava, Slovakia
2
Insitute of Virology, Biomedical Research Center, Slovak Academy of Sciences, Dubravska cesta 9, 845 05 Bratislava, Slovakia
3
Central European Institute of Technology, Masaryk University, 625 00 Brno, Czech Republic
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Department of Pediatrics, Medical Faculty of Masaryk University and University Hospital Brno, Černopolní 9, 625 00 Brno, Czech Republic
5
Institute for Clinical and Translational Research, Biomedical Research Center, Slovak Academy of Sciences, Dubravska cesta 9, 845 05 Bratislava, Slovakia
*
Author to whom correspondence should be addressed.
Genes 2019, 10(6), 459; https://doi.org/10.3390/genes10060459
Received: 19 May 2019 / Revised: 12 June 2019 / Accepted: 13 June 2019 / Published: 15 June 2019
(This article belongs to the Section Human Genomics and Genetic Diseases)
The molecular genetics of well-characterized inherited diseases, such as phenylketonuria (PKU) and hyperphenylalaninemia (HPA) predominantly caused by mutations in the phenylalanine hydroxylase (PAH) gene, is often complicated by the identification of many novel variants, often with no obvious impact on the associated disorder. To date, more than 1100 PAH variants have been identified of which a substantial portion have unknown clinical significance. In this work, we study the functionality of seven yet uncharacterized PAH missense variants p.Asn167Tyr, p.Thr200Asn, p.Asp229Gly, p.Gly239Ala, p.Phe263Ser, p.Ala342Pro, and p.Ile406Met first identified in the Czech PKU/HPA patients. From all tested variants, three of them, namely p.Asn167Tyr, p.Thr200Asn, and p.Ile406Met, exerted residual enzymatic activity in vitro similar to wild type (WT) PAH, however, when expressed in HepG2 cells, their protein level reached a maximum of 72.1% ± 4.9%, 11.2% ± 4.2%, and 36.6% ± 7.3% compared to WT PAH, respectively. Remaining variants were null with no enzyme activity and decreased protein levels in HepG2 cells. The chaperone-like effect of applied BH4 precursor increased protein level significantly for p.Asn167Tyr, p.Asp229Gly, p.Ala342Pro, and p.Ile406Met. Taken together, our results of functional characterization in combination with in silico prediction suggest that while p.Asn167Tyr, p.Thr200Asn, and p.Ile406Met PAH variants have a mild impact on the protein, p.Asp229Gly, p.Gly239Ala, p.Phe263Ser, and p.Ala342Pro severely affect protein structure and function. View Full-Text
Keywords: phenylalanine hydroxylase; phenylketonuria; BH4; functional studies; missense variants phenylalanine hydroxylase; phenylketonuria; BH4; functional studies; missense variants
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Pecimonova, M.; Kluckova, D.; Csicsay, F.; Reblova, K.; Krahulec, J.; Procházkova, D.; Skultety, L.; Kadasi, L.; Soltysova, A. Structural and Functional Impact of Seven Missense Variants of Phenylalanine Hydroxylase. Genes 2019, 10, 459.

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