Expression and maturation of human cathepsin D in baby-hamster kidney cells.
In medium and in homogenates from baby-hamster kidney cells (BHK) transfected with human cathepsin D cDNA, an elevated activity of cathepsin D was found as compared to non-transfected cells. The elevated activity was removed by titrating the homogenates with an anti-(human cathepsin D) antibody. Metabolic labelling and immunoprecipitation revealed that, in the transfected cells, human cathepsin D was synthesized as a 53-kDa precursor indistinguishable from that found in human cells. A portion of the precursor was secreted and the remainder was processed to intermediate and mature chains within a few hours of synthesis. The precursor that was released from the transfected cells had a slightly smaller apparent size than that from cultured human fibroblasts. This difference was abrogated when the precursors were treated with glycopeptidase F. In the intracellular small chain a difference was observed in the size of carbohydrate chains that were cleavable with endo-beta-N-acetylglucosaminidase H. Sequence analysis of the N-termini of mature intracellular cathepsin D indicated a N-terminal trimming in both large and small chains from both human and transfected hamster cells. The proteolytic maturation of human cathepsin D in BHK cells closely resembles that in human cells, whereas a portion of the carbohydrate side chains is processed differently. The trimming of the N-termini in mature cathepsin D is proposed to be a part of the maturation and aging of this protein.
Horst
M
M
Institut für Physiologische Chemie und Pathobiochemie, Westfälische Wilhelms-Universität, Münster, Germany.
Hasilik
A
A
D000118:Acetylglucosaminidase / Q000378:metabolism; D000581:Amidohydrolases; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002402:Cathepsin D / Q000235:genetics* / Q000276:immunology; D002478:Cells, Cultured; D006224:Cricetinae; D004247:DNA / Q000378:metabolism; D015870:Gene Expression; D014644:Genetic Variation; D006801:Humans; D006868:Hydrolysis; D007668:Kidney / Q000378:metabolism*; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011233:Precipitin Tests; D014162:Transfection
D000596:Amino Acids; D004247:DNA; D000118:Acetylglucosaminidase; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D002402:Cathepsin D; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase