Structural interpretation of mutations in phenylalanine hydroxylase protein aids in identifying genotype–phenotype correlations in phenylketonuria

Abstract

Phenylalanine hydroxylase (PAH) is the enzyme that converts phenylalanine to tyrosine as a rate-limiting step in phenylalanine catabolism and protein and neurotransmitter biosynthesis. Over 300 mutations have been identified in the gene encoding PAH that result in a deficient enzyme activity and lead to the disorders hyperphenylalaninaemia and phenylketonuria. The determination of the crystal structure of PAH now allows the determination of the structural basis of mutations resulting in PAH deficiency. We present an analysis of the structural basis of 120 mutations with a 'classified' biochemical phenotype and/or available in vitro expression data. We find that the mutations can be grouped into five structural categories, based on the distinct expected structural and functional effects of the mutations in each category. Missense mutations and small amino acid deletions are found in three categories: 'active site mutations', 'dimer interface mutations', and 'domain structure mutations'. Nonsense mutations and splicing mutations form the category of 'proteins with truncations and large deletions'. The final category, 'fusion proteins', is caused by frameshift mutations. We show that the structural information helps formulate some rules that will help predict the likely effects of unclassified and newly discovered mutations: proteins with truncations and large deletions, fusion proteins and active site mutations generally cause severe phenotypes; domain structure mutations and dimer interface mutations spread over a range of phenotypes, but domain structure mutations in the catalytic domain are more likely to be severe than domain structure mutations in the regulatory domain or dimer interface mutations. European Journal of Human Genetics (2000) 8, 683-696.

HowPAH gene mutations cause hyper-phenylalaninemia and why mechanism matters: Insights from in vitro expression

“…of a different amino acid residue to the properties of the residue that is normally present in the wild-type protein, it can be apparent that an important interaction between amino acids is lost, or that the replacement amino acid cannot be accommodated in a given space without seriously perturbing the surrounding environment. For catalogues of illustrative examples see Erlandsen and Stevens [1999] and Jennings et al [2000a].…”

“…As expected from the primary sequence, those amino acid residues that are altered as a result of disease-causing mutations are found to be distributed very widely throughout the structure. The 3D location and structural contacts of those individual amino acid residues have been documented [Erlandsen and Stevens, 1999;Jennings et al, 2000a]. In some cases, but not all, this can help illuminate the effects of a mutation upon protein function or structure, or generate hypotheses which may then be tested by IVE analysis.…”

Variations in genotype-phenotype correlations in phenylketonuria patients

“…The large number of possible allelic combinations makes prediction of the phenotype difficult (Dipple and McCabe, 2000;Scriver and Kaufman, 2001;Kasnauskiene et al, 2003;Kim et al, 2006;Bercovich et al, 2008;Danielle et al, 2009). In vitro expression analysis studies have demonstrated a large range of residual activities among different mutations (null to 75%) (Waters et al, 1998;Jennings et al, 2000;Waters, 2003).…”

Molecular Diagnosis of Phenylketonuria: From Defective Protein to Disease-Causing Gene Mutation

“…A structural model of phenylalanine hydro xylase was completed (14)(15)(16) and the location, along with structural contacts of individual amino acid residues, was documented (7,17). This provided the base for a molecular modeling of the mutation effect in silico (18).…”