title: Synthesis of related forms of the lysosomal enzyme alpha-mannosidase in Dictyostelium discoideum.
abstract: Purified lysosomal alpha-mannosidase from the cellular slime mold, Dictyostelium discoideum, is composed of two subunits of Mr = 58,000 and 60,000 as revealed by polyacrylamide gel electrophoresis under denaturing conditions. The pattern of peptide fragments produced when these two species are digested with proteases indicates that they are related but not identical. Using monoclonal antibodies prepared against purified alpha-mannosidase, we have analyzed the different forms of the enzyme synthesized in vivo. In addition to two bands that co-migrate with the pure enzyme, a large Mr (140,000) species is found in immunoprecipitated [35S] methionine-labeled extracts of cellular and secreted proteins. The precipitation of all three bands is inhibited by preadsorption of the antibodies with pure enzyme and all three proteins are absent in extracts of an alpha-mannosidase structural gene mutant. One-dimensional peptide maps indicate that all sequences present in the smaller species are found in the Mr = 140,000 form. In addition, the large form accounts for about 20% of the extracellular alpha-mannosidase activity secreted by amoebae during growth in axenic culture. These results lead to the conclusion that alpha-mannosidase is first synthesized as a large enzymatically active precursor which is modified and proteolytically cleaved to form the smaller subunits.
mesh_terms: D004023:Dictyostelium / Q000201:enzymology*; D004792:Enzyme Precursors / Q000378:metabolism; D008247:Lysosomes / Q000201:enzymology*; D008361:Mannosidases / Q000096:biosynthesis* / Q000302:isolation & purification; D008970:Molecular Weight; D043323:alpha-Mannosidase
chemical_list: D004792:Enzyme Precursors; D008361:Mannosidases; D043323:alpha-Mannosidase