Tetrahydrobiopterin Deficiency

Updated: Jan 27, 2016
  • Author: Noah S Scheinfeld, JD, MD, FAAD; Chief Editor: Luis O Rohena, MD, FAAP, FACMG  more...
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Tetrahydrobiopterin (BH4) deficiencies are disorders that affect phenylalanine (Phe or F) homeostasis, as well as brain biosynthesis of catecholamine, serotonin, and (occasionally) nitric oxide. [1]

In particular, excess production of reactive oxygen species causes oxidation of BH4, an essential cofactor. In cases of such stress, it possible that BH4, sepiapterin, folic acid resveratrol, and AVE3085 can have the ability to recouple endothelial nitric oxide synthase (eNOS) and improve endothelial function. [2] Mouse studies in the context of hypoxic pulmonary hypertension have assessed endothelial NO synthase expression and biopterin metabolism and eNOS expression, along with sepiapterin (a precursor of tetrahydrobiopterin) and eNOS gene removal. [3]

BH4 deficiencies are grouped with phenylketonuria (PKU), which is an inborn error of protein metabolism that results from an impaired ability to metabolize the essential amino acid Phe. Similar to PKU, BH4 deficiencies negatively affect developmental function; however, BH4 deficiencies also affect neurologic functioning, depending on the variant. Some, but not all, BH4 deficiencies may be detected with PKU screening tests used in Western countries, depending on the variant. The pathology of BH4 deficiencies explicates the types of oxidative stress that can also cause decreased BH4 from inherited hyperphenylalaninemia to mitochondrial diseases. [4]

BH4 deficiencies are heterogeneous. They range from mild forms that do not require treatment to severe cases that are difficult to ameliorate even with therapy.

Dihydrofolate reductase shields eNOS involving uncoupling in tetrahydrobiopterin deficiency. [5]

GTP-cyclohydrolase 1 (GCH1) and tetrahydrobiopterin are important for regulation of beta-adrenergic control of heart rate. [6]

Anxiety and depression in mice have been linked to tetrahydrobiopterin deficiency. [7]

Tetrahydrobiopterin is important for embryonic development. [8]



Enzymatic reactions and defects

BH4 deficiencies fall into 4 main categories, depending on the enzymatic defect that leads to a lack of BH4. Through September 2006, 193 different mutant alleles or molecular lesions identified in the guanosine triphosphate cyclohydrolase I (GCH1), 6-pyruvoyl-tetrahydropterin synthase (PTPS), sepiapterin reductase (SPR), carbinolamine-4a-dehydratase (PCD), or dihydropteridine reductase (DHPR) genes had been identified. [9]

The most well-established human function of BH4 is as the cofactor for Phe-4-hydroxylase (PAH), tyrosine-3-hydroxylase, and tryptophan-5-hydroxylase. The last 2 are key enzymes in biogenic amine biosynthesis, that is, aromatic amino acid synthesis. In addition to hydroxylating aromatic amino acids, BH4 serves as the cofactor for nitric oxide synthase and glyceryl-ether mono-oxygenase.

These reactions are based on the ability of BH4 to react with molecular oxygen to form an active oxygen intermediate that can hydroxylate substrates. Although BH4 is absolutely essential for nitric oxide synthase activity, its exact function with different forms of the enzyme and its mechanism of action remain to be defined.

BH4 is synthesized from guanosine triphosphate (GTP) in at least 4 enzymatic steps by the action of 3 enzymes. GTP cyclohydrolase I (GTPCH), the first enzyme in BH4 biosynthesis, catalyzes the formation of 7,8-dihydroneopterin triphosphate from GTP in a single reaction step. GTPCH is subject to feedback inhibition by BH4. A single-copy gene, GCH1, located on chromosome band 14q22.1-q22.2 encodes GTPCH.

In the next step, 6-pyruvoyl-tetrahydropterin synthase (PTPS) catalyzes the conversion of 7,8-dihydroneopterin triphosphate to 6-pyruvoyl-tetrahydropterin. The PTS gene on chromosome band 11q22.3-q23.3 encodes PTPS.

Sepiapterin reductase (SR) is a nicotinamide adenine dinucleotide phosphate (NADP), reduced form, (NADPH) oxidoreductase. It is required for the final 2-step reduction of the dike to intermediate 6-pyruvoyl-tetrahydropterin to BH4. The SPR gene on chromosome band 2p14-p12 encodes SR.

During the enzymatic hydroxylation of aromatic amino acids, molecular oxygen is consumed and BH4 is peroxidated and oxidized. The pterin intermediate is subsequently reduced back to BH4 by 2 enzymes and a reduced pyridine nucleotide (ie, NADH) in a complex recycling reaction.

Molecular oxygen is first bound to BH4 to form an unstable 4-alpha--peroxy-BH4. The mono-oxygenation of aromatic amino acids is thus concomitant with oxidation of BH4 to 4-alpha--hydroxy-BH4 (pterin-4-alpha-carbinolamine). Pterin-4-alpha-carbinolamine is subsequently dehydrated to quinonoid-dihydrobiopterin (q-dihydrobiopterin) and water by the specific and highly efficient pterin-4-alpha-carbinolamine dehydratase (PCD). The PCBD gene on chromosome band 10q22 encodes PCD.

In the last step of BH4 recycling, q-dihydrobiopterin is reduced back to BH4 by the NADH-DHPR. Folate inhibitors, such as methotrexate, inhibit the activity of the enzyme both in vivo and in vitro. The QHPR gene on chromosome band 4p15.3 encodes DHPR.

Genetic factors

BH4 deficiency comprises heterogeneous autosomal recessive disorders caused by mutations in the PTS (most common), SPR, QHPR, and GCH1 genes. See the image below for an illustration of autosomal recessive inheritance patterns.

Autosomal recessive inheritance. Autosomal recessive inheritance.

Defects in the SPR gene cause neurotransmitter deficiency without hyperphenylalaninemia (HPA); defects in the GTPCH gene (ie, GCH1) may also cause autosomal dominant dopa-responsive dystonia (DRD).

BH4 deficiency without HPA occurs in 6,10-methylenetetrahydrofolate reductase deficiency and vitiligo or DRD.

Fiori et al noted that HPA is an inherited metabolic disorder due to deficiency of the enzyme PAH or its cofactor BH4. [10] BH4 -responsiveness in PAH-deficient HPA is a recently described characteristic of most mild phenotypes. BH4 -responsive patients have reduced plasma Phe levels after the oral administration of BH4.

The investigators determined the incidence of BH4-responsiveness among a nonselected cohort of patients with PAH-deficient HPA and evaluated the phenotype-genotype correlations. They evaluated 11 patients born in Lombardy, Italy, between January 2000 and December 2004. HPA (107 patients) was classified after BH4 -loading test, an analysis of urinary pterin levels, and a determination of DHPR activity in the blood. The researchers assessed the patients for BH4 -responsiveness.

Patients were given 6R-BH4 20mg/kg orally as a single dose, and plasma samples were obtained at 0, 4, 8, and 24 hours after administration. In patients with basal plasma Phe levels of 360 mmol/L, a combined Phe (100 mg Phe/kg) and BH4 (20 mg/kg) loading test was performed. Patients were considered responsive to BH4 when their plasma Phe levels decreased by 30% at 8 hours after oral administration of BH4.

The investigators found that BH4 significantly lowered blood Phe levels in 91 (85%) of 107 patients with PAH-deficient HPA. Most of the responsive patients had mild HPA (77%), although some had mild (7%) or moderate (7%) PKU. One patient with classical PKU was responsive to BH BH4. Eighteen mutations were associated to the BH4 -responsive phenotype.

The authors concluded that a consistent number of patients with PAH-deficient HPA responded to BH4 and that BH4 responsiveness seemed to be common in mild phenotypes. Genotype was not the only factor that determined BH BH4 responsiveness.

Although dexamethasone reduces BH4 levels in endothelial cells, there is no evidence of eNOS uncoupling. The reduced NO production in- endothelial cells treated with dexamethasone is mainly attributable to reduced eNOS expression and decreased eNOS phosphorylation at serine 1177. [11]





United States

The incidence of classic PKU is approximately 1 case in 15,000 births. The incidence of BH4 deficiency is approximately 1 case per 1 million births, or 1.5-2% of cases of PKU.


In Taiwan, 2-30% of cases of PKU are attributed to BH4 deficiency. [12] In Turkey, which has the highest incidence of PKU in the world with approximately 1 case per 2600 births, 15% of cases are due to BH4 deficiency. In Saudi Arabia, 66% of PKU cases are due to BH4 deficiency. The incidence also appears to be increased in southern Brazil. [13] Such increased incidences are thought to be related to consanguinity.

Pangkanon et al reported the first 2 cases of PTPS deficiency in Thailand. [14] Both cases were males with phenylalanine levels of 25.23 mg/dL and 23.4 mg/dL, respectively. The urinary pterins analysis showed low biopterin levels, low percentages of urinary biopterin, and high neopterin levels. The mutation analysis of the patient with a phenylalanine level of 25.23 mg/dL revealed a point mutation in exon 4 and a homozygous C-to-T transition at nucleotide 200 in codon 67 (T67M). The other patient demonstrated a compound heterozygous in exon 4, C-to-A transition at nucleotide 200 and exon 5, and C-to-T transition at nucleotide 259 of the PTS gene, confirming PTPS deficiency.

Farrugia et al noted that the deficient activity of the DHPR enzyme is due to mutations in the quinoid dihydropteridine reductase (QDPR) gene on 4p15.3. [15] Deficient activity of the DHPR enzyme results in defective recycling of BH4, and homozygotes have a rare form of atypical HPA and PKU. The heterozygote frequency in the Maltese population is high (3.3%).

A multicenter study from China reported 256 cases of BH4 deficiency, with a majority (59.9%) residing in Eastern China. [16] In 93% of patients, 6-pyruvoyl-tetrahydropterin synthase (PTPS) deficiency was the resultant cause of BH4 deficiency. Four hotspot mutations produced 76.6 % of PTS genetic mutations. Researcher identified 2 new variants in the QDPR gene. Eighty-three percent of patients underwent diet therapy, L-dopa, BH4, and/or 5-hydroxytryptophan treatment. While the WISC IQ score average was 80, 62.7% of patients had normal scores. Researchers observed a normal range of physical development and few adverse treatment effects (7.8%)


Patients with severe BH4 deficiency present with mental retardation and neurologic impairment. Early death may result. Patients with mild cases can have mild degrees of mental retardation and neurologic impairment.

Treatment can significantly alter the course of disease. A Japanese study [17] of 19 patients who received sapropterin dihydrochloride treatment along with L-dopa and 5-hydroxytryptophan for BH4 deficiency before age 4 years were followed for up to 28 years and had excellent results.

Similar findings showing the impact of prompt therapy were noted in a study of 626 patients with different genetic causes for their BH4 deficiency, although the exact genetic mutation and the differing times from diagnosis to treatment make the outcome for BH4 deficiencies highly variable. [18]


Children of Chinese, Turkish, and Saudi Arabian descent are most often affected.


No sex predilection is reported. The mode of inheritance is autosomal recessive.


BH4 deficiency is most commonly diagnosed in newborns by means of newborn screening programs. Consider BH4 deficiency in patients of any age who have PKU and developmental delay or mental retardation with neurologic impairment. Newborn screening does not always detect the disease. Patients who are symptomatic usually present by age 4 months.