Identification of two novel β-mannosidosis-associated sequence variants: Biochemical analysis of β-mannosidase (MANBA) missense mutations

β-Mannosidosis (OMIM # 248510) is an autosomal-recessive lysosomal storage disorder caused by deficiency of the lysosomal enzyme β-mannosidase (MANBA, E.C. 3.2.1.25). The disorder has been reported in goat, cattle and man. The human disorder is rare and only 20 cases in 16 families have been reported. We have sequenced the exons and exon-intron borders in a European patient with infantile onset of β-mannosidosis. The patient was compound heterozygous for a silent mutation (c.375A > G) in exon 3 causing alternative splicing, and a missense mutation (c.1513T > C, p.Ser505Pro) in exon 12. The alternative splicing event deleted four nucleotides from the transcript and was predicted to result in premature termination of translation. In order to evaluate the consequence of the missense mutation, we inserted the human β-mannosidase gene into an expression vector, performed site-directed mutagenesis and expressed the normal and mutant enzyme in COS-7 cells. We also included the previously reported β-mannosidosis-associated missense mutations c.544C > T (p.Arg182Trp) and c.1175G > A (p.Gly392Glu), which were found in patients presenting a milder phenotype. Cells transfected with the wild-type construct showed a 33-fold increase in β-mannosidase activity compared to mock-transfected cells, whereas cells transfected with the mutant constructs showed no detectable increase in activity. We propose that the milder phenotype described in some β-mannosidosis patients with missense mutations in the MANBA gene is not due to residual β-mannosidase activity, but rather caused by epigenetic and/or environmental factors.