Molecular characterization and gene disruption of mouse lysosomal putative serine carboxypeptidase 1.
The retinoid-inducible serine carboxypeptidase 1 (Scpep1; formerly RISC) is a lysosomal matrix protein that was initially identified in a screen for genes induced by retinoic acid. Recently, it has been spotlighted by several proteome analyses of the lysosomal compartment, but its cellular function and properties remain unknown to date. In this study, Scpep1 from mice was analysed with regard to its intracellular processing into a mature dimer consisting of a 35 kDa N-terminal fragment and a so far unknown 18 kDa C-terminal fragment and the glycosylation status of the mature Scpep1 fragment. Although Scpep1 shares notable homology and a number of structural hallmarks with the well-described lysosomal carboxypeptidase protective protein/cathepsin A, the purified recombinant 55 kDa precursor and the homogenates of Scpep1-overexpressing cells do not show proteolytic activity or increased serine carboxypeptidase activity towards artificial serine carboxypeptidase substrates. Hence, we disrupted the Scpep1 gene in mice by a gene trap cassette, resulting in a Scpep1/beta-galactosidase/neomycin phosphotransferase fusion protein. The fusion protein is devoid of the C-terminal half of Scpep1, including two amino acids of the assumed catalytic triad which is indispensable for its predicted serine carboxypeptidase activity. However, Scpep1-deficient mice were viable and fertile, and did not exhibit either lysosomal storage or reduced lysosomal SC activity under any tested condition.
Kollmann
Katrin
K
Abteilung Biochemie II, Georg-August Universität Göttingen, Göttingen, Germany.
Damme
Markus
M
Deuschl
Florian
F
Kahle
Jörg
J
D'Hooge
Rudi
R
Lüllmann-Rauch
Renate
R
Lübke
Torben
T
D000595:Amino Acid Sequence; D000818:Animals; D002268:Carboxypeptidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D006031:Glycosylation; D008247:Lysosomes / Q000201:enzymology* / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016415:Sequence Alignment
D002268:Carboxypeptidases; C486671:Scpep1 protein, mouse