Large Neutral Amino Acid Supplementation Exerts Its Effect through Three Synergistic Mechanisms: Proof of Principle in Phenylketonuria Mice

Vliet, Danique van; Bruinenberg, Vibeke M.; Mazzola, Priscila Nicolao; Faassen, Martijn H. J. R. van; Blaauw, Pim de; Heiner‐Fokkema, M. Rebecca; Anholt, R. D. Van; Zee, Eddy A. van der; Spronsen, Francjan J. van

doi:10.1371/journal.pone.0143833

Cited by 66 publications

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“…Both strains showed a vast increase in Phe levels and disrupted serotonin and norepinephrine levels in the brain (for norepinephrine a trend was observed for BTBR statistically) which is in accordance with previous studies (Andolina et al, 2011; Pascucci et al, 2013; Sawin et al, 2014; van Vliet et al, 2015). How these PKU-related changes can result in a different functional outcome is not clear.…”

Section: Discussionsupporting

confidence: 91%

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The Behavioral Consequence of Phenylketonuria in Mice Depends on the Genetic Background

Bruinenberg

Goot

Vliet

et al. 2016

Front. Behav. Neurosci.

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To unravel the role of gene mutations in the healthy and the diseased state, countless studies have tried to link genotype with phenotype. However, over the years, it became clear that the strain of mice can influence these results. Nevertheless, identical gene mutations in different strains are often still considered equals. An example of this, is the research done in phenylketonuria (PKU), an inheritable metabolic disorder. In this field, a PKU mouse model (either on a BTBR or C57Bl/6 background) is often used to examine underlying mechanisms of the disease and/or new treatment strategies. Both strains have a point mutation in the gene coding for the enzyme phenylalanine hydroxylase which causes toxic concentrations of the amino acid phenylalanine in blood and brain, as found in PKU patients. Although the mutation is identical and therefore assumed to equally affect physiology and behavior in both strains, no studies directly compared the two genetic backgrounds to test this assumption. Therefore, this study compared the BTBR and C57Bl/6 wild-type and PKU mice on PKU-relevant amino acid- and neurotransmitter-levels and at a behavioral level. The behavioral paradigms were selected from previous literature on the PKU mouse model and address four domains, namely (1) activity levels, (2) motor performance, (3) anxiety and/or depression-like behavior, and (4) learning and memory. The results of this study showed comparable biochemical changes in phenylalanine and neurotransmitter concentrations. In contrast, clear differences in behavioral outcome between the strains in all four above-mentioned domains were found, most notably in the learning and memory domain. The outcome in this domain seem to be primarily due to factors inherent to the genetic background of the mouse and much less by differences in PKU-specific biochemical parameters in blood and brain. The difference in behavioral outcome between PKU of both strains emphasizes that the consequence of the PAH mutation is influenced by other factors than Phe levels alone. Therefore, future research should consider these differences when choosing one of the genetic strains to investigate the pathophysiological mechanism underlying PKU-related behavior, especially when combined with new treatment strategies.

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Section: Discussionsupporting

confidence: 91%

“…Plasma and brain samples were stored at −80°C until further processing. Amino acid and neurotransmitter analyses were performed as described in van Vliet et al (2015). …”

Section: Methodsmentioning

confidence: 99%

The Behavioral Consequence of Phenylketonuria in Mice Depends on the Genetic Background

Bruinenberg

Goot

Vliet

et al. 2016

Front. Behav. Neurosci.

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“…A Phe-mentes LNAA-keveréket tartalmazó készítmények orális úton történő alkalmazása egyrészt csökkentheti a vér Phe-szintet azáltal, hogy kompetitív módon gátolja a Phe bélből történő felszívódását, másrészt a vér-agy gáton átjutó Phe csökkentésével, valamint a cerebralis Tyr-és Trp-szintek növelésével javítja az aminosav-egyensúlyt a központi idegrendszerben [36][37][38][39]. Alkalmazása szigorú körültekintést igényel.…”

Section: Diétás Terápiákunclassified

Phenylketonuria felnőttkorban

2017

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A phenylketonuria 1975 óta az újszülöttkori tömegszűrés része. Mára már hazánkban is felnőtt egy olyan generáció, amely születésétől kezdve speciális diétát tart és orvosi tápszert fogyaszt. A szűrés és a korai terápia bevezetésének köszönhetően a phenylketonuriás gyerekek normális életvitellel bíró felnőtté válhatnak. Ennek köszönhetően a phenylketonuria többé már nem egy gyermekgyógyászati betegség. Ebből adódóan a felnőtt phenylketonuriások ellátása anyagcsere-specializációval bíró belgyógyászati centrumokban történik. Az élethosszig tartó speciális terápia, a phenylketonuria felnőttkori vonatkozásait most ismerjük meg. Az összefoglaló közlemény célja bemutatni a phenylketonuriával kapcsolatos újdonságokat, fókuszálva a felnőttkori vonatkozásokra. Hosszú évek után nemzetközi irány-elvek jelentek meg, új terápiás lehetőségek váltak elérhetővé és várhatóak a közeljövőben, új kihívásokkal kell szembenéznünk, mint a maternalis phenylketonuria, a dietoterápia hosszú távú kihatásai vagy a nem kezelt phenylketonuriás felnőttek szövődményei. Orv Hetil. 2017; 158(47): 1857-1863. Kulcsszavak: phenylketonuria, felnőtt, metabolikus kontroll Adult phenylketonuriaStarting from 1975 phenylketonuria is part of the newborn screening program in Hungary. Since then a generation, treated with special diet and medical foods right after neonatal diagnosis has reached adulthood. Thanks to early treatment initiation, children with phenylketonuria are able to lead life to the full. Consequently, phenylketonuria is no longer considered a pediatric disease. Follow up of adult patients with phenylketonuria is performed in internal medicine centers specialized in metabolic diseases. The outcome of the lifelong special treatment, and the particularities of phenylketonuria in adulthood are yet to be determined. The aim of our review is to present recent findings in phenylketonuria focusing mainly on the adult care. After long time the first international guidelines appeared, new therapies were put in use, and these current developments are expected to be implemented in daily practice in the near future. New challenges must be met such as maternal phenylketonuria, long term effects of dietotherapy and the sequelae of untreated phenylketonuria in adulthood. Rövidítések AAVV = (adeno-associated viral vector) adenoasszociált virális vektor; BH 4 = (tetrahydrobiopterin) tetrahydrobiopterin; BMD = (bone mineral density) csontsűrűség; ETPKU = (earlytreated phenylketonuria) korán kezelt phenylketonuria; GMP = (glycomacropeptide) glikomakropeptid; LNAA = (large neutral amino acids) nagy neutrális aminosavak; MRI = (magnetic resonance imaging) mágneses rezonanciás vizsgálat; PAH = (phenylalanine hydroxylase) fenilalanin-hidroxiláz; PAL = (phenylalanine ammonia-lyase) fenilalanin-ammónia-liáz; Phe = (phenylalanine) fenilalanin; PKU = (phenylketonuria) phenylketonuria; QOL = (quality of life) életminőség; Trp = (tryptophan) triptofán; TYH = (tyrosine hydroxylase) tirozinhidroxiláz; Tyr = (tyrosine) tirozin; WMA = (white matter abnormalities) fehérállo...

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“…This event increases the level of Phe in the brain in PKU, leading to various complications (Pardridge, 1998). One way of achieving a low level of Phe in blood and brain is by the intake of high levels of LNAA (Soltanizadeh & Mirmoghtadaie, 2014;van Vliet et al, 2015van Vliet et al, , 2018van Vliet et al, , 2019. Supplementation of LNAA decreases the uptake of Phe and consequently decreases the level of Phe in the brain, as a result of competition for the common transporter in the gastrointestinal tract and BBB.…”

Section: Introductionmentioning

confidence: 99%

Improvement, cloning, and expression of an in silico designed protein enriched with large neutral amino acids in Pichia pastoris for possible application in phenylketonuria

Appaiah

Vasu

2020

J Food Biochem

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Phenylketonuria (PKU) is an inborn disease caused by defective phenylalanine hydroxylase, which consequently results in the accumulation of phenylalanine in the brain leading to further complications. One of the promising approaches in dietary treatment is the supplementation of large neutral amino acid (LNAA). The LNAA compete with phenylalanine for the common L‐type LNAA transporter across the blood–brain barrier, and decrease phenylalanine levels in the brain. In this study, the earlier LNAA‐enriched protein model was improved (Protein Model‐66) and validated in silico. The reverse translated and codon‐optimized synthetic LNAA66 gene was cloned into pPICZαC and expressed in Pichia pastoris. The expressed protein was purified by His Select affinity chromatography. SDS‐PAGE and Western blotting analysis showed a band at an expected molecular weight of 12 kDa, confirming the expression of the modeled protein. To our knowledge, this is the first report showing the cloning and expression of an in silico designed LNAA‐enriched protein. Practical applications One of the promising dietary treatment of phenylketonuria (PKU) is the supplementation of large neutral amino acid (LNAA), wherein high levels of LNAA compete with phenylalanine for the same L‐type LNAA transporter, and consequently decrease phenylalanine accumulation in the brain, thereby decreasing neurological complications. For the first time, here, we are showing that an in silico designed and validated Protein Model‐66, rich in LNAA, can be successfully cloned and expressed in Pichia pastoris. The complete biochemical and structural characterization of this protein will give a clear insight into its potential application for PKU treatment. The protein can be potentially used as a supplement to treat PKU to those who are non‐adherent to the restricted, non‐palatable, and expensive diet. Furthermore, this novel and effective strategy of in silico designing, cloning and expression can be exploited to develop proteins for various applications of industrial, food, medical, and academic relevance.

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Large Neutral Amino Acid Supplementation Exerts Its Effect through Three Synergistic Mechanisms: Proof of Principle in Phenylketonuria Mice

Cited by 66 publications

References 44 publications

The Behavioral Consequence of Phenylketonuria in Mice Depends on the Genetic Background

The Behavioral Consequence of Phenylketonuria in Mice Depends on the Genetic Background

Phenylketonuria felnőttkorban

Improvement, cloning, and expression of an in silico designed protein enriched with large neutral amino acids in Pichia pastoris for possible application in phenylketonuria

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