title: Proprotein processing within secretory dense core granules of Tetrahymena thermophila.
abstract: In the ciliate Tetrahymena thermophila, the polypeptides stored in secretory dense core granules (DCGs) are generated by proteolytic processing of precursors, and the mature products assemble as a crystal. Previous observations suggested that this maturation involves precise cleavage at distinct motifs by a small number of enzymes. To test these inferences, we analyzed the determinants for site-specific processing of pro-Grl1p (Granule lattice protein 1) by complete gene replacement with modified alleles. Contrary to the predictions of previous models, none of the component amino acids in a putative processing motif was necessary for targeted cleavage. Indeed, cleavage at a range of alternative positions near the native site was consistent with normal DCG assembly. Furthermore, substitution of other classes of processing site motifs did not perturb DCG structure or function. These results suggest that processing can be catalyzed by multiple proteases, for which substrate accessibility may be the prime determinant of site specificity. Consistent with this, inhibition of either subtilisin or cathepsin family proteases resulted in delayed processing of pro-Grl1p.
authors.affiliation: Department of Molecular Genetics and Cell Biology, the University of Chicago, Illinois 60637, USA.
authors.forename: Niels R
authors.initials: NR
authors.lastname: Bradshaw
authors.forename: N Doane
authors.initials: ND
authors.lastname: Chilcoat
authors.forename: John W
authors.initials: JW
authors.lastname: Verbsky
authors.forename: Aaron P
authors.initials: AP
authors.lastname: Turkewitz
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011498:Protein Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017386:Sequence Homology, Amino Acid; D016808:Tetrahymena thermophila / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: D011498:Protein Precursors