Activity and processing of wild type and glycosylation site mutated PPT1. (A) COS-1 cells were transfected with plasmids producing mutated PPT1 enzymes and lysed 48 h post-transfection. PPT1 enzyme activity of each lysate (5 μg total protein) was analyzed by a one-hour reaction in + 37°C. Transfection efficacies were controlled by immunofluorescence analysis. The data shown represents an average of three independent experiments and the standard deviations between these experiments are marked as error bars. (B) COS-1 cells were transfected and metabolically labelled with 35S-Cysteine for 1 hour, chased for 2 hours, immunoprecipitated, separated in SDS-PAGE and autoradiographed. Both intracellular and secreted forms of PPT are shown.