Toward mechanistic models for genotype-phenotype correlations in phenylketonuria using protein stability calculations.


Phenylketonuria (PKU) is a genetic disorder caused by variants in the gene encoding phenylalanine hydroxylase (PAH), resulting in accumulation of phenylalanine to neurotoxic levels. Here, we analyzed the cellular stability, localization, and interaction with wild-type PAH of 20 selected PKU-linked PAH protein missense variants. Several were present at reduced levels in human cells, and the levels increased in the presence of a proteasome inhibitor, indicating that proteins are proteasome targets. We found that all the tested PAH variants retained their ability to associate with wild-type PAH, and none formed aggregates, suggesting that they are only mildly destabilized in structure. In all cases, PAH variants were stabilized by the cofactor tetrahydrobiopterin (BH4 ), a molecule known to alleviate symptoms in certain PKU patients. Biophysical calculations on all possible single-site missense variants using the full-length structure of PAH revealed a strong correlation between the predicted protein stability and the observed stability in cells. This observation rationalizes previously observed correlations between predicted loss of protein destabilization and disease severity, a correlation that we also observed using new calculations. We thus propose that many disease-linked PAH variants are structurally destabilized, which in turn leads to proteasomal degradation and insufficient amounts of cellular PAH protein.

Submission Details


Submitter: Shu-Ching Ou

Submission Date: March 22, 2019, 11:40 a.m.

Version: 1

Publication Details
Scheller R;Stein A;Nielsen SV;Marin FI;Gerdes AM;Di Marco M;Papaleo E;Lindorff-Larsen K;Hartmann-Petersen R,Hum Mutat (2019) Toward mechanistic models for genotype-phenotype correlations in phenylketonuria using protein stability calculations. PMID:30648773
Additional Information

Study Summary

Number of data points 8002
Proteins Phenylalanine-4-hydroxylase
Unique complexes 7981
Assays/Quantities/Protocols Computational Protocol: ΔΔG ; Computational Protocol: Enzyme activity (% of WT)
Libraries Variants for PAH

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
399.2 A,B,C,D Phenylalanine-4-hydroxylase P04176 PH4H_RAT
394.0 A,B,C,D Phenylalanine-4-hydroxylase P16331 PH4H_MOUSE
370.0 A,B,C,D Phenylalanine-4-hydroxylase P00439 PH4H_HUMAN
364.8 A,B,C,D Phenylalanine-4-hydroxylase Q2KIH7 PH4H_BOVIN