Phenylketonuria (PKU), caused by variants in the phenylalanine hydroxylase (PAH) gene, is the most common autosomal-recessive Mendelian phenotype of amino acid metabolism. We estimated that globally 0.45 million individuals have PKU, with global prevalence 1:23,930 live births (range 1:4,500 [Italy]-1:125,000 [Japan]). Comparing genotypes and metabolic phenotypes from 16,092 affected subjects revealed differences in disease severity in 51 countries from 17 world regions, with the global phenotype distribution of 62% classic PKU, 22% mild PKU, and 16% mild hyperphenylalaninemia. A gradient in genotype and phenotype distribution exists across Europe, from classic PKU in the east to mild PKU in the southwest and mild hyperphenylalaninemia in the south. The c.1241A>G (p.Tyr414Cys)-associated genotype can be traced from Northern to Western Europe, from Sweden via Norway, to Denmark, to the Netherlands. The frequency of classic PKU increases from Europe (56%) via Middle East (71%) to Australia (80%). Of 758 PAH variants, c.1222C>T (p.Arg408Trp) (22.2%), c.1066-11G>A (IVS10-11G>A) (6.4%), and c.782G>A (p.Arg261Gln) (5.5%) were most common and responsible for two prevalent genotypes: p.[Arg408Trp];[Arg408Trp] (11.4%) and c.[1066-11G>A];[1066-11G>A] (2.6%). Most genotypes (73%) were compound heterozygous, 27% were homozygous, and 55% of 3,659 different genotypes occurred in only a single individual. PAH variants were scored using an allelic phenotype value and correlated with pre-treatment blood phenylalanine concentrations (n = 6,115) and tetrahydrobiopterin loading test results (n = 4,381), enabling prediction of both a genotype-based phenotype (88%) and tetrahydrobiopterin responsiveness (83%). This study shows that large genotype databases enable accurate phenotype prediction, allowing appropriate targeting of therapies to optimize clinical outcome.

• Klíčová slova
• BH4, PAH deficiency, PKU, hyperphenylalaninemia, phenylalanine, tetrahydrobiopterin,
• MeSH
• alely MeSH
• biopteriny analogy a deriváty   genetika   MeSH
• fenotyp MeSH
• fenylalanin krev   MeSH
• fenylalaninhydroxylasa genetika   MeSH
• fenylketonurie krev   epidemiologie   genetika   MeSH
• frekvence genu genetika   MeSH
• genetická predispozice k nemoci genetika   MeSH
• genetické asociační studie metody   MeSH
• genotyp MeSH
• homozygot MeSH
• lidé MeSH
• mutace genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Geografické názvy
• Evropa MeSH
• Názvy látek
• biopteriny MeSH
• fenylalanin MeSH
• fenylalaninhydroxylasa MeSH
• sapropterin MeSH Prohlížeč

Mutations can be induced by environmental factors but also arise spontaneously during DNA replication or due to deamination of methylated cytosines at CpG dinucleotides. Sites where mutations occur with higher frequency than would be expected by chance are termed hotspots while sites that contain mutations rarely are termed coldspots. Mutations are permanently scanned and repaired by repair systems. Among them, the mismatch repair targets base pair mismatches, which are discriminated from canonical base pairs by probing altered elasticity of DNA. Using biased molecular dynamics simulations, we investigated the elasticity of coldspots and hotspots motifs detected in human genes associated with inherited disorders, and also of motifs with Czech population hotspots and de novo mutations. Main attention was paid to mutations leading to G/T and A+/C pairs. We observed that hotspots without CpG/CpHpG sequences are less flexible than coldspots, which indicates that flexible sequences are more effectively repaired. In contrary, hotspots with CpG/CpHpG sequences exhibited increased flexibility as coldspots. Their mutability is more likely related to spontaneous deamination of methylated cytosines leading to C > T mutations, which are primarily targeted by base excision repair. We corroborated conclusions based on computer simulations by measuring melting curves of hotspots and coldspots containing G/T mismatch.

• Klíčová slova
• DNA bending, Muts protein, free energy calculations, hotspots–coldspots, mutations,
• MeSH
• CpG ostrůvky MeSH
• DNA chemie   genetika   MeSH
• lidé MeSH
• mutace * MeSH
• nukleotidové motivy * MeSH
• simulace molekulární dynamiky * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• DNA MeSH

The molecular genetics of well-characterized inherited diseases, such as phenylketonuria (PKU) and hyperphenylalaninemia (HPA) predominantly caused by mutations in the phenylalanine hydroxylase (PAH) gene, is often complicated by the identification of many novel variants, often with no obvious impact on the associated disorder. To date, more than 1100 PAH variants have been identified of which a substantial portion have unknown clinical significance. In this work, we study the functionality of seven yet uncharacterized PAH missense variants p.Asn167Tyr, p.Thr200Asn, p.Asp229Gly, p.Gly239Ala, p.Phe263Ser, p.Ala342Pro, and p.Ile406Met first identified in the Czech PKU/HPA patients. From all tested variants, three of them, namely p.Asn167Tyr, p.Thr200Asn, and p.Ile406Met, exerted residual enzymatic activity in vitro similar to wild type (WT) PAH, however, when expressed in HepG2 cells, their protein level reached a maximum of 72.1% ± 4.9%, 11.2% ± 4.2%, and 36.6% ± 7.3% compared to WT PAH, respectively. Remaining variants were null with no enzyme activity and decreased protein levels in HepG2 cells. The chaperone-like effect of applied BH4 precursor increased protein level significantly for p.Asn167Tyr, p.Asp229Gly, p.Ala342Pro, and p.Ile406Met. Taken together, our results of functional characterization in combination with in silico prediction suggest that while p.Asn167Tyr, p.Thr200Asn, and p.Ile406Met PAH variants have a mild impact on the protein, p.Asp229Gly, p.Gly239Ala, p.Phe263Ser, and p.Ala342Pro severely affect protein structure and function.

• Klíčová slova
• BH4, functional studies, missense variants, phenylalanine hydroxylase, phenylketonuria,
• MeSH
• biopteriny analogy a deriváty   chemie   genetika   MeSH
• buňky Hep G2 MeSH
• fenylalaninhydroxylasa chemie   genetika   MeSH
• fenylketonurie genetika   metabolismus   patologie   MeSH
• genotyp MeSH
• lidé MeSH
• missense mutace genetika   MeSH
• počítačová simulace MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• biopteriny MeSH
• fenylalaninhydroxylasa MeSH
• sapropterin MeSH Prohlížeč

Inherited ichthyoses belong to a large and heterogeneous group of mendelian disorders of cornification, and can be distinguished by the quality and distribution of scaling and hyperkeratosis, by other dermatologic and extracutaneous involvement, and by inheritance. We present the genetic analysis results of probands with X-linked ichthyosis, autosomal recessive congenital ichthyosis, keratinopathic ichthyosis, and a patient with Netherton syndrome. Genetic diagnostics was complemented by in silico missense variant analysis based on 3D protein structures and commonly used prediction programs to compare the yields of these two approaches to each other. This analysis revealed various structural defects in proteins coded by mutated genes while no defects were associated with known polymorphisms. Two patients with pathogenic variants in the ABCA12 gene have a premature termination codon mutation on one allele and a silent variant on the second. The silent variants c.69G > A and c.4977G > A are localised in the last nucleotide of exon 1 and exon 32, respectively, and probably affect mRNA splicing. The phenotype of both patients is very severe, including a picture harlequin foetus after birth; later (at 3 and 6 years of age, respectively) ectropin, eclabion, generalised large polygonal scaling and erythema.

• Klíčová slova
• 3D protein structure, Autosomal recessive congenital ichthyosis, In silico analysis, Keratinopathic ichthyosis,
• MeSH
• ABC transportéry genetika   MeSH
• fenotyp MeSH
• genetická predispozice k nemoci genetika   MeSH
• ichtyóza etiologie   genetika   MeSH
• lidé MeSH
• nesmyslný kodon genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• dopisy MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• ABC transportéry MeSH
• ABCA12 protein, human MeSH Prohlížeč
• nesmyslný kodon MeSH

Hyperphenylalaninemia (HPA) is one of the most common metabolic disorders. HPA, which is transmitted by an autosomal recessive mode of inheritance, is caused by mutations of the phenylalanine hydroxylase gene. Most mutations are missense and lead to reduced protein stability and/or impaired catalytic function. The impact of such mutations varies, ranging from classical phenylketonuria (PKU), mild PKU, to non-PKU HPA phenotypes. Despite the fact that HPA is a monogenic disease, clinical data show that one PKU genotype can be associated with more in vivo phenotypes, which indicates the role of other (still unknown) factors. To better understand the phenotype-genotype relationships, we analyzed computationally the impact of missense mutations in homozygotes stored in the BIOPKU database. A total of 34 selected homozygous genotypes was divided into two main groups according to their phenotypes: (A) genotypes leading to non-PKU HPA or combined phenotype non-PKU HPA/mild PKU and (B) genotypes leading to classical PKU, mild PKU or combined phenotype mild PKU/classical PKU. Combining in silico analysis and molecular dynamics simulations (in total 3 μs) we described the structural impact of the mutations, which allowed us to separate 32 out of 34 mutations between groups A and B. Testing the simulation conditions revealed that the outcome of mutant simulations can be modulated by the ionic strength. We also employed programs SNPs3D, Polyphen-2, and SIFT but based on the predictions performed we were not able to discriminate mutations with mild and severe PKU phenotypes.

• MeSH
• fenotyp MeSH
• fenylalaninhydroxylasa chemie   genetika   MeSH
• fenylketonurie genetika   patologie   MeSH
• genotyp MeSH
• konformace proteinů MeSH
• lidé MeSH
• missense mutace genetika   MeSH
• počítačová simulace MeSH
• simulace molekulární dynamiky MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fenylalaninhydroxylasa MeSH

Myotonia congenita (MC) is a genetic disease caused by mutations in the skeletal muscle chloride channel gene (CLCN1) encoding the skeletal muscle chloride channel (ClC-1). Mutations of CLCN1 result in either autosomal dominant MC (Thomsen disease) or autosomal recessive MC (Becker disease). The ClC-1 protein is a homodimer with a separate ion pore within each monomer. Mutations causing recessive myotonia most likely affect properties of only the mutant monomer in the heterodimer, leaving the wild type monomer unaffected, while mutations causing dominant myotonia affect properties of both subunits in the heterodimer. Our study addresses two points: 1) molecular genetic diagnostics of MC by analysis of the CLCN1 gene and 2) structural analysis of mutations in the homology model of the human dimeric ClC-1 protein. In the first part, 34 different types of CLCN1 mutations were identified in 51 MC probands (14 mutations were new). In the second part, on the basis of the homology model we identified the amino acids which forming the dimer interface and those which form the Cl(-) ion pathway. In the literature, we searched for mutations of these amino acids for which functional analyses were performed to assess the correlation between localisation of a mutation and occurrence of a dominant-negative effect (corresponding to dominant MC). This revealed that both types of mutations, with and without a dominant-negative effect, are localised at the dimer interface while solely mutations without a dominant-negative effect occur inside the chloride channel. This work is complemented by structural analysis of the homology model which provides elucidation of the effects of mutations, including a description of impacts of newly detected missense mutations.

• MeSH
• chloridové kanály chemie   genetika   metabolismus   MeSH
• dospělí MeSH
• fenotyp MeSH
• konformace proteinů MeSH
• kosterní svaly metabolismus   MeSH
• lidé MeSH
• missense mutace MeSH
• mladiství MeSH
• mladý dospělý MeSH
• molekulární modely MeSH
• multimerizace proteinu MeSH
• mutace * MeSH
• myotonia congenita diagnóza   genetika   MeSH
• Check Tag
• dospělí MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• chloridové kanály MeSH
• CLC-1 channel MeSH Prohlížeč