Identifying and elucidating the roles of Y198N and Y204F mutations in the PAH enzyme through molecular dynamic simulations

Abstract

Phenylketonuria is an autosomal recessive disorder caused by mutations in the phenylalanine hydroxylase gene. In phenylketonuria causes various symptoms including severe mental retardation. PAH gene of a classical Phenylketonuria patient was sequenced, and two novel heterozygous mutations, p.Y198N and p.Y204F, were found. This study aimed to reveal the impacts of these variants on the structural stability of the PAH enzyme. In-silico analyses using prediction tools and molecular dynamics simulations were performed. Mutations were introduced to the wild type catalytic monomer and full length tetramer crystal structures. Variant pathogenicity analyses predicted p.Y198N to be damaging, and p.Y204F to be benign by some prediction tools and damaging by others. Simulations suggested p.Y198N mutation cause significant fluctuations in the spatial organization of two catalytic residues in the temperature accelerated MD simulations with the monomer and increased root-mean-square deviations in the tetramer structure. p.Y204F causes noticeable changes in the spatial positioning of T278 suggesting a possible segregation from the catalytic site in temperature accelerated MD simulations with the monomer. This mutation also leads to increased root-mean-square fluctuations in the regulatory domain which may lead to conformational change resulting in inhibition of dimerization and enzyme activation. Our study reports two novel mutations in the PAH gene and gives insight to their effects on the PAH activity. MD simulations did not yield conclusive results that explains the phenotype but gave plausible insight to possible effects which should be investigated further with in-silico and in-vitro studies to assess the roles of these mutations in etiology of PKU.

Communicated by Ramaswamy H. Sarma

Keywords:

• Phenylketonuria
• phenylalanine hydroxylase
• PAH
• molecular dynamics simulation
• hyperphenylalaninemia
• NAMD
• VMD

Disclosure statement

No potential conflict of interest was reported by the authors.

Availability of data and materials

All of the data and materials used in this study are included in this article and its supplementary files.

Authors’ contributions

TA performed DNA isolation from patient sample, analysis of Sanger sequencing. TA, AYK and UG performed all remaining experiments, prepared figures and tables, and drafted the manuscript. AÇAZ and EK diagnosed the patient, measured blood Phe and Tyr levels and provided clinical data. TA, AYK, OUS and NBI designed the study and contributed to data analysis. All authors read and approved the final version of the manuscript.

Consent for publication

Not applicable.

Ethics approval and consent to participate

The study was approved by the Bogazici University Institutional Review Board for Research with Human Subjects (2016/31), and all patients or their guardians gave written informed consent.

Additional information

Funding

This study was funded by Bogazici University Scientific Research Projects (11480). Funding body had no role in the design of the study, data collection, analysis, interpretation of data, writing the manuscript or decision to publish.