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Toward mechanistic models for genotype–phenotype correlations in phenylketonuria using protein stability calculations
Author(s) -
Scheller Rasmus,
Stein Amelie,
Nielsen Sofie V.,
Marin Frederikke I.,
Gerdes AnneMarie,
Marco Miriam,
Papaleo Elena,
LindorffLarsen Kresten,
HartmannPetersen Rasmus
Publication year - 2019
Publication title -
human mutation
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 162
eISSN - 1098-1004
pISSN - 1059-7794
DOI - 10.1002/humu.23707
Subject(s) - missense mutation , phenylalanine hydroxylase , tetrahydrobiopterin , phenotype , proteasome , phenylalanine , biology , hyperphenylalaninemia , protein stability , genotype , gene , genetics , biochemistry , cofactor , enzyme , amino acid
Abstract 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 (BH 4 ), 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.