Synthesis and processing of arylsulfatase A in human skin fibroblasts

Biosynthesis of arylsulfatase A in normal and mutant human fibroblasts was studied by growing cells in the presence of L-[4,5-3H] leucine or [2-3H] mannose, isolation of labelled arylsulfatase A by immune precipitation and visualization of electrophoretically separated polypeptide by fluorography. Arylsulfatase A was synthesized as a precursor with a mean apparent molecular mass of 62 kDa. Intracellularly the precursor was converted into a 60.5 kDa polypeptide within a chase period of 1 to 7 days. The 60.5 kDa product in polyacrylamide corresponded to one of two polypeptides present in arylsulfatase A isolated from human placenta. In fibroblasts from a patient with metachromatic leukodystrophy no immune precipitable polypeptides of arylsulfatase A were detected. In normal fibroblasts less than 10% of the precursor of arylsulfatase A was secreted into the medium, whereas in mucolipidosis II fibroblasts and in control fibroblasts grown in the presence of NH4Cl up to 90% of the precursor of arylsulfatase A, appeared in the medium and remained there without change in the apparent molecular mass for at least 7 days. Arylsulfatase A polypeptides appear to contain two carbohydrate side chains. In about 90% of the polypeptides both side chains are cleaved by endo-beta-N-acetylglucosaminidase H, whereas in the remaining chains one of the two oligosaccharides is not cleaved.