Analysis of Phenylalanine Hydroxylase Genotypes and Hyperphenylalaninemia Phenotypes Using L-[1-13C]Phenylalanine Oxidation Rates in Vivo: A Pilot Study1

Treacy, Eileen P.; Delente, Jacques; Elkas, G; Carter, K; Lambert, Marie; Waters, Paula J.; Scriver, Charles R.

doi:10.1203/00006450-199710000-00002

Cited by 41 publications

(36 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, since the in vitro PAH activity is proportional to in vivo PAH activity in hepatocytes, as demonstrated by the correlation of genotype and biochemical phenotype, the predicted PAH activity system is useful for assessing the relative severity of PKU mutations, the classification of PKU, and the prediction of the clinical course. Secondly, there are cases with discordance between genotype and biochemical phenotype (Kayaalp et al 1997;Treacy et al 1997). Treacy et al (1996) have demonstrated different in vivo metabolic profiles for phenylalanine arising from differences in phenylalanine transamination and renal clearance of transamination metabolites in two siblings with a different PKU phenotype but the same genotype.…”

Section: Discussionmentioning

confidence: 97%

Molecular characterization of phenylketonuria in Japanese patients

Okano¹,

Asada²,

Kang³

et al. 1998

Human Genetics

View full text Add to dashboard Cite

We characterized phenylalanine hydroxylase (PAH) genotypes of Japanese patients with phenylketonuria (PKU) and hyperphenylalaninemia (HPA). PKU and HPA mutations in 41 Japanese patients were identified by denaturing gradient gel electrophoresis and direct sequencing, followed by restriction fragment length polymorphism analysis to find a large deletion involving exons 5 and 6. Of 82 mutant alleles, 76 (92%) were genotyped showing 21 mutations. The major mutations were R413P (30.5%), R243Q (7.3%), R241 C (7.3%), IVS4nt-1 (7.3%), T2781 (7.3%), E6nt-96A-->g (6.1%), Y356X (4.9%), R111X (3.7%), and 442-706delE5/6 (2.4%). Eight new mutations (L52 S, delS70, S70P, Y77X, IVS3nt-1, A132 V, W187 C, and C265Y) and a polymorphism of IVS10nt-14 were detected. In vitro PAH activities of mutant PAH cDNA constructs were determined by a COS cell expression system. Six mutations, viz., R408Q, L52 S, R241 C, S70P, V388 M, and R243Q, had 55%, 27%, 25%, 20%, 16% and 10% of the in vitro PAH activity of normal constructs, respectively. The mean pretreatment phenylalanine concentration (0.83+/-0.21 mmol/l) of patients carrying the R408Q, R241 C, or L52 S mutation and a null mutation was significantly lower (P<0.0005) than that (1.99+/-0.65 mmol/l) of patients with both alleles carrying mutations associated with a severe genotype. Simple linear regression analysis showed a correlation between pretreatment phenylalanine concentrations and predicted PAH activity in 29 Japanese PKU patients (y=31.9-1.03x, r=0.59, P<0.0001). Genotype determination is useful in the prediction of biochemical and clinical phenotypes in PKU and can be of particular help in managing patients with this disorder.

show abstract

Section: Discussionmentioning

confidence: 97%

Molecular characterization of phenylketonuria in Japanese patients

Okano¹,

Asada²,

Kang³

et al. 1998

Human Genetics

View full text Add to dashboard Cite

show abstract

“…Neither RBC gal-1-P nor urine galactitol increased within 24 h of testing. Other metabolic disorders that express a serious problem in newborns have been studied following bolus or sustained exposure to trace amount of precursors in blocked reactions (18,19). Children with maple syrup urine disease and phenylketonuria had no adverse events to bolus or sustained infusions of leucine or phenylalanine respectively at these trace doses for 13 C detection in breath.…”

Section: Discussionmentioning

confidence: 99%

Screening Newborns for Galactosemia Using Total Body Galactose Oxidation to CO2 in Expired Air

Barbouth

Velazquez

Konopka³

et al. 2007

Pediatr Res

View full text Add to dashboard Cite

Classic galactosemia is caused by impaired galactose-1-phosphate uridyltransferase (GALT EC 2.7.712). If discovered and treated within the first days of life, the acute problems of hepatocellular damage, sepsis, and death are prevented. However, chronic problems such as ataxia, tremor, dyspraxic speech, and ovarian failure may occur. To determine whether screening newborns before discharge from the nursery for GALT deficiency is feasible and whether acute and chronic signs could be prevented by earlier intervention, we developed a simplified "breath test." We quantitated total body oxidation of 13 C-D-galactose to 13 CO 2 in expired air by normal newborns between 2 h and 2 mo of age and compared their results to older children with GALT deficiency. We found no differences in total body galactose oxidation (TBGO) among normal newborns up to 48 h of age, but a 2-fold rise in TBGO developed during their first 2 wk of life. Older children with galactosemia had significantly less oxidative capacity than normal newborns. We conclude that newborn breath testing for total body galactose oxidation is feasible before discharge from nursery. It has potential utility for both preventing acute neonatal toxicity and determining the mechanisms producing long-term complications such as ovarian failure, dyspraxia, ataxia, and tremors. C lassic galactosemia is an autosomal recessive disorder caused by a deficiency of galactose-1-phosphate uridyltransferase (GALT EC 2.7.7.12). The biochemical phenotype is characterized by accumulation of galactose, galactose-1-phosphate, and galactitol in cells, and of excess excretion of galactitol and galactose (1-4). The disease incidence is approximately 1:30,000 live births in the United States (2). If untreated in the first 2 wk of life, poor feeding, jaundice, hepatic failure, Escherichia coli sepsis, and death may occur. We have personal experience with families whose newborns had died (http://www.savebabies.org/diseasedescriptions/galactosemia. php, formally known as, "Tyler for Life Foundation" and personal experience).Despite population-based newborn screening and dietary intervention within 2-3 wk of life, the clinical outcome of patients with classic galactosemia is variable. Long-term complications include premature ovarian failure, dyspraxic speech, short stature, learning disabilities, and cataracts (5-7). In one ex post facto study, outcome was not related to time of treatment (5).Most states in the United States screen for galactosemia by the Beutler Method, which measures GALT in dried blood on filter paper through enzyme-linked fluorescence of NADPH production from glucose-1-phosphate. Some states use total galactose quantization (8,9). The median time for results to be received by a referring physician from public health laboratories is poorly documented nationally but from personal experience varies between 4 and 30 d (9).It is not clear whether this prolonged exposure to excess galactose-1-phosphate produces the chronic effects of galactosemia during fetal developm...

show abstract

“…The R157N MBP-PAH band was repeatedly reduced in intensity after Coomassie staining of SDS-PAGE gels, despite the loading of apparently equal amounts of protein as determined by the Bradford assay. Normal absolute values are provided in Treacy et al, (1997). e Not detectable; threshold is 3% of normal.…”

Section: Expression Of Mbp-pah Fusion Proteins In E Colimentioning

confidence: 99%

Alterations in protein aggregation and degradation due to mild and severe missense mutations (A104D, R157N) in the human phenylalanine hydroxylase gene (PAH)

et al. 1998

View full text Add to dashboard Cite

Phenylalanine hydroxylase (PAH) catalyzes the conversion of phenylalanine to tyrosine; its activity is the major determinant of phenylalanine disposal. Mutations in the corresponding human gene (PAH), which encodes the human hepatic PAH enzyme, result in hyperphenylalaninemia; the resulting phenotypes can range in severity from mild forms of hyperphenylalaninemia with benign outcome to the severe form, phenylketonuria with impaired cognitive development. This paper describes the detailed characterization of two inherited recessive missense mutations in PAH, c.311C→A (A104D) and [c.470G→A;c.471A→C] (R157N), which are associated, respectively, in the homozygous or functionally hemizygous states, with mild and severe metabolic phenotypes. We used three different in vitro PAH expression systems (in Escherichia coli, cell‐free rabbit reticulocyte lysates, and human embryonal kidney cells), as well as a unique assay for phenylalanine oxidation in vivo. In each system, we observed alterations of PAH function and physical properties, compared with wild‐type enzyme, and differences in relative severity of effects between these two mutations. Pulse‐chase experiments showed increased PAH degradation, probably related to observed aberrations in protein folding and altered oligomerization, as a basic mechanism underlying effects of these missense mutations. Hum Mutat 12:344–354, 1998.© 1998 Wiley‐Liss, Inc.

show abstract

Analysis of Phenylalanine Hydroxylase Genotypes and Hyperphenylalaninemia Phenotypes Using L-[1-13C]Phenylalanine Oxidation Rates in Vivo: A Pilot Study1

Cited by 41 publications

References 24 publications

Molecular characterization of phenylketonuria in Japanese patients

Molecular characterization of phenylketonuria in Japanese patients

Screening Newborns for Galactosemia Using Total Body Galactose Oxidation to CO2 in Expired Air

Alterations in protein aggregation and degradation due to mild and severe missense mutations (A104D, R157N) in the human phenylalanine hydroxylase gene (PAH)

Contact Info

Product

Resources

About