Biosynthesis and maturation of glucocerebrosidase in Gaucher fibroblasts.
The biosynthesis and maturation of glucocerebrosidase were studied in fibroblasts from patients with the neurological and non-neurological forms of Gaucher disease and in control cells. In control fibroblasts the precursor of glucocerebrosidase (62-63 kDa), observed after a short pulse with [35S]methionine, was converted during the chase period to a 66-kDa intermediate form and, finally, to the 59-kDa mature protein. In fibroblasts from patients with the non-neurological phenotype of Gaucher disease (type 1) the same biosynthetic forms were seen as in control fibroblasts. These biosynthetic forms correspond to the three-banded pattern seen in control and Gaucher type 1 fibroblast extracts analysed by the immunoblotting procedure, or after electrophoresis and fluorography of extracts of such fibroblasts cultured for 5 days with [14C]leucine. The 59-kDa protein seen in type 1 fibroblasts was unstable and disappeared after a prolonged chase; this disappearance was not observed when the cells were grown in the presence of leupeptin. In fibroblasts from patients with the neurological forms of Gaucher disease (types 2 and 3) the 62.5-kDa precursor of glucocerebrosidase was present in near-normal amounts after a short pulse, but the 59-kDa form was not detected even when cells were cultured with leupeptin. These results are in accordance with the absence of the 59-kDa band in immunoblots of types 2 and 3 fibroblast extracts. Culturing of type 1, type 2 and type 3 Gaucher fibroblasts in the presence of leupeptin led to an increase in the activity of glucocerebrosidase.
D002460:Cell Line; D003429:Cross Reactions; D005347:Fibroblasts / Q000201:enzymology; D005776:Gaucher Disease / Q000201:enzymology*; D005959:Glucosidases / Q000096:biosynthesis*; D005962:Glucosylceramidase / Q000096:biosynthesis*; D006801:Humans; D007163:Immunosorbent Techniques; D007976:Leupeptins / Q000494:pharmacology; D008715:Methionine / Q000378:metabolism; D008970:Molecular Weight; D009154:Mutation
D007976:Leupeptins; D008715:Methionine; D005959:Glucosidases; D005962:Glucosylceramidase; C032854:leupeptin