Gaucher disease: single gene molecular characterization of one-hundred Indian patients reveals novel variants and the most prevalent mutation

Abstract

Background: Gaucher disease is a rare pan-ethnic, lysosomal storage disorder resulting due to beta-Glucosidase (GBA1) gene defect. This leads to the glucocerebrosidase enzyme deficiency and an increased accumulation of undegraded glycolipid glucocerebroside inside the cells' lysosomes. To date, nearly 460 mutations have been described in the GBA1 gene. With the aim to determine mutations spectrum and molecular pathology of Gaucher disease in India, the present study investigated one hundred unrelated patients (age range: 1 day to 31 years) having splenomegaly, with or without hepatomegaly, cytopenia and bone abnormality in some of the patients.

Methods: The biochemical investigation for the plasma chitotriosidase enzyme activity and β-Glucosidase enzyme activity confirmed the Gaucher disease. The mutations were identified by screening the patients' whole GBA gene coding region using bidirectional Sanger sequencing.

Results: The biochemical analysis revealed a significant reduction in the β-Glucosidase activity in all patients. Sanger sequencing established 71 patients with homozygous mutation and 22 patients with compound heterozygous mutation in GBA1 gene. Lack of identification of mutations in three patients suggests the possibility of either large deletion/duplication or deep intronic variations in the GBA1 gene. In four cases, where the proband died due to confirmed Gaucher disease, the parents were found to be a carrier. Overall, the study identified 33 mutations in 100 patients that also covers four missense mutations (p.Ser136Leu, p.Leu279Val, p.Gly383Asp, p.Gly399Arg) not previously reported in Gaucher disease patients. The mutation p.Leu483Pro was identified as the most commonly occurring Gaucher disease mutation in the study (62% patients). The second common mutations identified were p.Arg535Cys (7% patients) and RecNcil (7% patients). Another complex mutation Complex C was identified in a compound heterozygous status (3% patients). The homology modeling of the novel mutations suggested the destabilization of the GBA protein structure due to conformational changes.

Conclusions: The study reports four novel and 29 known mutations identified in the GBA1 gene in one-hundred Gaucher patients. The given study establishes p.Leu483Pro as the most prevalent mutation in the Indian patients with type 1 Gaucher disease that provide new insight into the molecular basis of Gaucher Disease in India.

Keywords: Chitotriosidase; GBA1 gene; Gaucher disease; Glucocerebrosidase; Indian population; novel mutations in GBA1 gene; p.Leu483Pro most common mutation; β-Glucosidase.

Fig. 1

Illustrative representation of the distributions of the variants identified in Indian Gaucher patients investigated in this study. a Majority of the patients were affected with Gaucher disease type I (77 patients), followed by Gaucher disease type II (12 patients) and Gaucher disease type III (11 patients). b The most common mutation c.1448T>C was observed in 62 patients (including homozygotes and compound heterozygotes). The second most common mutations identified were c.1603C>T and RecNcil. c Variations on exon 4–12 were observed on GBA1 gene. Also a mutation g.3548A>G was observed in the intron 1. The mutations were more clustered on exon 4 and exon 7 to exon 11

Fig. 2

Identification of novel variants in GBA1 gene. a Illustrative representation of the distributions of the novel variants identified in Indian Gaucher patients investigated in this study. b Sanger sequencing discovered four missense variants in GBA1 gene. The variants p.Ser136Leu, p.Gly383Asp, and p.Gly399Arg, were identified, as compound heterozygotes along with another know mutant allele. The variant p.Leu279Val existed in homozygous form. An arrow indicates the point of variation. c The multiple alignment of the protein sequence surrounding the novel variants against various orthologous sequence revealed the conservative status of the wildtype residues (marked red)

Fig. 3

Homology modeling of novel missense variants identified in the GBA1 gene. The native structure (blue) and mutant structure (brown) are superimposed. a The model of the variant p.Ser136Leu depicting the changes in the loop region joining α1 and α1a; at the codon number 136 (TCA-TTA). b The model of the variant p.Leu279Val depicting the destabilization of the normal shape of the active site cavity at the codon number 279 (CTG-GTG). c The model of the variant p.Gly383Asp depicting the conformational changes in the loop regions at the codon number 383 (GGC-GAC). d The model of the variant p.Gly399Arg depicting the conformational changes in the alpha helix at the codon number 399 (GGG-CGG). All the models reveal the conformational changes in the GBA protein structure