The lysosomal degradation of sphingolipids with short oligosaccharide chains depends on small glycosylated non-enzymatic sphingolipid activator proteins (SAPs, saposins). Four of the five known SAPs, SAP-A, -B, -C and -D, are derived by proteolytic processing from a common precursor protein (SAP-precursor) that is encoded by a gene on chromosome 10 consisting of 15 exons and 14 introns. SAP-B is a non-specific glycolipid binding protein that stimulates in vitro the hydrolysis of about 20 glycolipids by different enzymes. In vivo SAP-B stimulates in particular the degradation of sulphatides by arylsulphatase A. So far, four different point mutations have been identified on the SAP-B domain of the SAP-precursor gene. The mutations result in a loss of mature SAP-B, causing the lysosomal accumulation of sulphatides and other sphingolipids, resulting in variant forms of metachromatic leukodystrophy (MLD). Here we report on a patient with SAP-B deficiency that is caused by a new homoallelic point mutation that has been identified by mRNA and DNA analysis. A 643A > C transversion results in the exchange of asparagine 215 to histidine and eliminates the single glycosylation site of SAP-B. Metabolic labelling experiments showed that the mutation had no effect on the intracellular transport of the encoded precursor to the acidic compartments and its maturation in the patient's cells. All four SAPs (SAP-A to SAP-D) were detectable by immunochemical methods. SAP-B in the patient's cells was found to be slightly less stable than the protein in normal cells and corresponded in size to the deglycosylated form of the wild-type SAP-B. Feeding studies with non-glycosylated SAP-precursor, generating non-glycosylated SAP-B, showed that the loss of the carbohydrate chain reduced the intracellular activity of the protein significantly. The additional structural change of the patient's SAP-B, caused by the change of amino acid 215 from asparagine to histidine, presumably resulted in an almost completely inactive protein.