Novel mutations in 3-phosphoglycerate dehydrogenase (PHGDH) are distributed throughout the protein and result in altered enzyme kinetics

Abstract

Three-phosphoglycerate dehydrogenase (3-PGDH) deficiency is a rare recessive inborn error in the biosynthesis of the amino acid L-serine characterized clinically by congenital microcephaly, psychomotor retardation, and intractable seizures. The biochemical abnormalities associated with this disorder are low concentrations of L-serine, D-serine, and glycine in cerebrospinal fluid (CSF). Only two missense mutations (p.V425M and p.V490M) have been identified in PHGDH, the gene encoding 3-PGDH, but it is currently unclear how these mutations in the carboxy-terminal regulatory domain of the protein affect enzyme function. We now describe five novel mutations in five patients with 3-PGDH deficiency; one frameshift mutation (p.G238fsX), and four missense mutations (p.R135W, p.V261M, p.A373T, and p.G377S). The missense mutations were located in the nucleotide binding and regulatory domains of 3-PGDH and did not affect steady-state expression, protein stability, and protein degradation rates. Patients' fibroblasts displayed a significant, but incomplete, reduction in maximal enzyme activities associated with all missense mutations. In transient overexpression studies in HEK293T cells, the p.A373T, p.V425M, and p.V490M mutations resulted in almost undetectable enzyme activities. Molecular modeling of the p.R135W and p.V261M mutations onto the partial crystal structure of 3-PGDH predicted that these mutations affect substrate and cofactor binding. This prediction was confirmed by the results of kinetic measurements in fibroblasts and transiently transfected HEK293T cells, which revealed a markedly decreased V(max) and an increase in K(m) values, respectively. Taken together, these data suggest that missense mutations associated with 3-PGDH deficiency either primarily affect substrate binding or result in very low residual enzymatic activity.

Expanding the clinical spectrum of 3‐phosphoglycerate dehydrogenase deficiency

“…Enzymatic analysis of 3-PGDH activity in cultured skin fibroblasts displayed a deficient value in both patients (6.8 and 3.1 respectively normal values 33.1 ± 1.9 (nmol/min.mg protein)), results that were comparable to what was reported in patients with the severe infantile phenotype of 3-PGDH deficiency (Tabatabaie et al 2009). Mutation analysis of the PHGDH gene confirmed a homozygous c.1129 G > A (p.G377S) mutation in both children.…”

“…Their plasma and CSF serine values were in the same range as what was observed in severely affected infants. The same was true for the 3-PGDH enzyme activity in cultured skin fibroblasts showing a residual activity comparable to that in patients with the infantile phenotype (Tabatabaie et al 2009). It is obvious that the mild clinical phenotype could not be predicted from the biochemical or molecular analysis and that additional genetic or environmental factors considerably modulate disease severity in serine deficiency.…”

“…Both parents were shown to be carriers of the mutation confirming autosomal recessive inheritance. The details of the molecular analysis were published elsewhere (Tabatabaie et al 2009). …”

Mutations inSLC1A4, encoding the brain serine transporter, are associated with developmental delay, microcephaly and hypomyelination

“…In spite of the poor permeability of L-serine through the blood–brain barrier, treatment of patients suffering from L-serine biosynthetic defects with L-serine seemed beneficial, alleviating seizure severity and frequency 6 14…”

Clinical review of genetic epileptic encephalopathies

“…3-PGD is inherited in an autosomal recessive manner, and diagnosis can be confirmed by evaluation of enzymatic activity or analysis of the PHGDH gene. A small number of patients have been reported with varying mutations along the PHGDH gene [112,113]. Oral l -serine treatment (up to 600 mg/kg/ day in six divided doses) has shown beneficial biochemical and clinical effects [105].…”