Identification of mutations that causes glucose-6-phosphate transporter defect in tunisian patients with glycogenosis type 1b

Abstract

Background: Glycogen storage disease type 1b (GSD1b) is an autosomal recessive lysosomal storage disease caused by defective glucose-6-phosphate transporter encoded by SLC37A4 leading to the accumulation of glycogen in various tissues. The high rate of consanguineous marriages in Tunisian population provides an ideal environment to facilitate the identification of homozygous pathogenic mutations. We aimed to determine the clinical and genetic profiles of patients with GSD1b to evaluate SLC37A4 mutations spectrum in Tunisian patients.

Methods: All exons and flanking intron regions of SLC37A4 gene were screened by direct sequencing to identify mutations and polymorphisms in three unrelated families with GSD1b. Bioinformatics tools were then used to predict the impacts of identified mutations on the structure and function of protein in order to propose a function-structure relationship of the G6PT1 protein.

Results: Three patients (MT, MB and SI) in Families I, II and III who had the severe phenotype were homoallelic for the two identified mutations: p.R300H (famillies I, II) and p.W393X (Family III), respectively. One of the alterations was a missense mutation p.R300H of exon 6 in SLC37A4 gene. The analysis of the protein structure flexibility upon p.R300H mutation using DynaMut tool and CABS-flex 2.0 server showed that the reported mutation increase the molecule flexibility of in the cytosol region and would probably lead to significant conformational changes.

Conclusion: This is the first Tunisian report of SLC37A4 mutations identified in Tunisia causing the glycogenosis type Ib disease. Bioinformatics analysis allowed us to establish an approximate structure-function relationship for the G6PT1 protein, thereby providing better genotype/phenotype correlation knowledge.

Keywords: Bioinformatics tool; Glucose-6-phosphate transporter (SLC37A4); Glycogenosis; Mutations.

Fig. 1

Family pedigree and electropherograms showing the reported mutation p.R300H and p.W393X associated with Glycogen storage disease type 1b (a) The right and left panel indicate the sequence electropherogram of the SLC37A4 mutations identified in exon 6 and 9 in the patients with GSD1b, respectively. (b) The right, the middle and the left panel concern the family pedigree of the patient MT, MB, and SI, respectively

Fig. 2

The Crystallographic structure analysis of the generated G6PT 3D structure model A) The analysis of the interactomic interaction before (left panel) and after mutations (right panel) introduction is indicated in the upper row. Wild-type Arg300 and mutant residues His300 are indicated dotted arrow and colored in pink and green, respectively. They showed as sticks alongside with the surrounding amino acids which are implicated on any type of interactions. B) The analysis of Electrostatic potentials before (left panel) and after mutations introduction (right panel) is shown in the middle and bottom rows, respectively. In this work, Electrostatic potentials were calculated using ionic strengths corresponding to 0 mM ion concentration and εP = 4. The negative and positive electrostatic potentials are highlighted in red and blue, respectively. C) The variation on protein stability after mutation introduction using DynaMut is shown in the bottom row. The amino acids are colored according to the vibrational entropy change upon mutation. Blue represents a rigidification of the structure and red a gain in flexibility. Backbone RMSD per residue for the wild type (red) and mutated protein (blue) are indicated in red and blue, respectively