In vitro expression analysis of R68G and R68S mutations in phenylalanine hydroxylase gene.

Zekanowsk, C; Pérez, Belén; Desviat, Lourdes R.; Wiszniewski, Wojciech; Ugarte, Magdalena

doi:10.18388/abp.2000_4016

Cited by 8 publications

(2 citation statements)

References 12 publications

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“…1A), since the predicted effect on protein stability ( kcal/ DDG p 8.7 mol) is abnormally high with respect to its very high in vitro residual activity (table 1) and mild folding effect. 57 Arg68 is located at the subunit interface in the dimer, 58 and a high DDG value for the R68G mutant appears to be partly explained by a relatively high contribution of the subunit binding energy to the change in stability (see the "Effects of Subunit Binding Energies in Oligomer Stability" subsection). Other mutations for which inconsistencies are encountered using different models are marked in table 5.…”

Section: Correlation Between In Vitro Residual Activity and Energeticmentioning

confidence: 99%

Predicted Effects of Missense Mutations on Native-State Stability Account for Phenotypic Outcome in Phenylketonuria, a Paradigm of Misfolding Diseases

Pey

Stricher

Serrano

et al. 2007

The American Journal of Human Genetics

165

145

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Phenylketonuria (PKU) is a genetic disease caused by mutations in human phenylalanine hydroxylase (PAH). Most missense mutations result in misfolding of PAH, increased protein turnover, and a loss of enzymatic function. We studied the prediction of the energetic impact on PAH native-state stability of 318 PKU-associated missense mutations, using the protein-design algorithm FoldX. For the 80 mutations for which expression analyses have been performed in eukaryote systems, in most cases we found substantial overall correlations between the mutational energetic impact and both in vitro residual activities and patient metabolic phenotype. This finding confirmed that the decrease in protein stability is the main molecular pathogenic mechanism in PKU and the determinant for phenotypic outcome. Metabolic phenotypes have been shown to be better predicted than in vitro residual activities, probably because of greater stringency in the phenotyping process. Finally, all the remaining 238 PKU missense mutations compiled at the PAH locus knowledgebase (PAHdb) were analyzed, and their phenotypic outcomes were predicted on the basis of the energetic impact provided by FoldX. Residues in exons 7-9 and in interdomain regions within the subunit appear to play an important structural role and constitute hotspots for destabilization. FoldX analysis will be useful for predicting the phenotype associated with rare or new mutations detected in patients with PKU. However, additional factors must be considered that may contribute to the patient phenotype, such as possible effects on catalysis and interindividual differences in physiological and metabolic processes.

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Section: Correlation Between In Vitro Residual Activity and Energeticmentioning

confidence: 99%

Predicted Effects of Missense Mutations on Native-State Stability Account for Phenotypic Outcome in Phenylketonuria, a Paradigm of Misfolding Diseases

Pey

Stricher

Serrano

et al. 2007

The American Journal of Human Genetics

165

145

View full text Add to dashboard Cite

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“…Thanks to the implementation of PKU screening, there are today thousands of individuals who, due to a timely diagnosis and subsequent treatment, can fulfill their intellectual and physical potentials. Although newborn screening has been in use for over 40 years in Poland, there exists a group of adults whose multiple health problems related to remote consequences of neurotoxic phenylalanine (Phe) effects occur, due to a lack of or a delay in medical intervention (Rolling, 1984;Zekanowski et al, 2000;Kasim et al, 2001;Giżewska, 2006). To date, very few coordinated actions have been undertaken in Poland in order to detect this particular group of patients (Giżewska, 2006).…”

Section: Introductionmentioning

confidence: 99%

Measurement of functional independence level and falls-risk in individuals with undiagnosed phenylketonuria.

Mazur¹,

Jarochowicz²,

Ołtarzewski³

et al. 2009

Acta Biochim Pol

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The aim of the study was to determine the level of functional independence in adult patients with previously undiagnosed or untreated phenylketonuria (PKU). The study was conducted among 400 intellectually impaired adult residents of Social Welfare Homes in South-Eastern Poland born prior to the introduction of neonatal PKU screening programs. PKU was screened by filter paper test using tandem mass spectrometry methods, and confirmed by gas chromatography-mass spectrometric analysis of PKU organic acids in urine. Degree of functional independence included the assessment of activities of daily living (Barthel Index) and measures of balance and gait (Tinetti scale). Eleven individuals with previously untreated PKU were identified whereby eight presented with moderate disability and three with mild disability. Six had a high risk of falls and five had a moderate risk of falls. This study indicates that there is considerable number of undiagnosed PKU patients within the Polish population who require assessment and management in order to reduce the impact of the neurological and neuropsychiatric problems associated with the condition. Appropriate therapy for those with undiagnosed PKU should, in particular, address the risk of falls.

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Protein stability and in vivo concentration of missense mutations in phenylalanine hydroxylase

2011

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A previous computational analysis of missense mutations linked to monogenic disease found a high proportion of missense mutations affect protein stability, rather than other aspects of protein structure and function. The purpose of the present study is to relate the presence of such stability damaging missense mutations to the levels of a particular protein present under ‘in vivo’ like conditions, and to test the reliability of the computational methods. Experimental data on a set of missense mutations of the enzyme phenylalanine hydroxylase (PAH) associated with the monogenic disease phenylketonuria (PKU) have been compared with the expected in vivo impact on protein function, obtained using SNPs3D, an in silico analysis package. A high proportion of the PAH mutations are predicted to be destabilizing. The overall agreement between predicted stability impact and experimental evidence for lower protein levels is in accordance with the estimated error rates of the methods. For these mutations, destabilization of protein three dimensional structure is the major molecular mechanism leading to PKU, and results in a substantial reduction of in vivo PAH protein concentration. Although of limited scale, the results support the view that destabilization is the most common mechanism by which missense mutations cause monogenic disease. In turn, this conclusion suggests the general therapeutic strategy of developing drugs targeted at restoring wild type stability.

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In vitro expression analysis of R68G and R68S mutations in phenylalanine hydroxylase gene.

Cited by 8 publications

References 12 publications

Predicted Effects of Missense Mutations on Native-State Stability Account for Phenotypic Outcome in Phenylketonuria, a Paradigm of Misfolding Diseases

Predicted Effects of Missense Mutations on Native-State Stability Account for Phenotypic Outcome in Phenylketonuria, a Paradigm of Misfolding Diseases

Measurement of functional independence level and falls-risk in individuals with undiagnosed phenylketonuria.

Protein stability and in vivo concentration of missense mutations in phenylalanine hydroxylase

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