Acid sphingomyelinase (ASM) is a lysosomal
hydrolase that degrades
sphingomyelin into
ceramide and
phosphocholine. Recent crystallographic studies revealed the functional role of the N-terminal ASM
saposin domain.
ASM deficiency due to mutations in the ASM-encoding
sphingomyelin phosphodiesterase 1 (SMPD1) gene causes an autosomal recessive
sphingolipid-storage disorder, known as
Niemann-Pick disease Type A (NPA) or Type B (
NPB). NPA is an early-onset neuronopathic disorder, while
NPB is a late-onset non-neuronopathic disorder. A homozygous one-base substitution (c.398G>A) of the SMPD1 gene was identified in an infant with NPA, diagnosed with complete loss of ASM activity in the patient's fibroblasts. This mutation is predicted to substitute
tyrosine for
cysteine at
amino acid residue 133, abbreviated as p.C133Y. The patient showed developmental delay, hepatosplenomegaly and rapid neurological deterioration leading to death at the age of 3 years. To characterize p.C133Y, which may disrupt one of the three
disulfide bonds of the N-terminal ASM
saposin domain, we performed immunoblotting analysis to explore the expression of a mutant ASM
protein in the patient's fibroblasts, showing that the
protein was detected as a 70-kDa
protein, similar to the wild-type ASM
protein. Furthermore, transient expression of p.C133Y ASM
protein in COS-7 cells indicated complete loss of ASM
enzyme activity, despite that the p.C133Y ASM
protein was properly localized to the lysosomes. These results suggest that the proper three-dimensional structure of
saposin domain may be essential for ASM catalytic activity. Thus, p.C133Y is associated with complete loss of ASM activity even with stable
protein expression and proper subcellular localization.