Formation of enzymatically active, homotypic, and heterotypic tetramers ofmouse mast cell tryptases - Dependence on a conserved Trp-rich domain on the surface

Mouse mast cell protease (mMCP) 6 and mMCP-7 are homologous tryptases store d in granules as macromolecular complexes with heparin and/or chondroitin s ulfate E containing serglycin proteoglycans, When pro-mMCP-7 and pseudozymo gen forms of this tryptase and mMCP-6 were separately expressed in insect c ells, all three recombinant proteins were secreted into the conditioned med ium as properly folded, enzymatically inactive 33-kDa monomers, However, wh en their propeptides were removed, mMCP 6 and mMCP-7 became enzymatically a ctive and spontaneously assumed an similar to 150-kDa tetramer structure. H eparin was not required for this structural change. When incubated at 37 de grees C, recombinant mMCP-7 progressively lost its enzymatic activity in a time-dependent manner. Its N-linked glycans helped regulate the thermal sta bility of mMCP-7. However, the ability of this tryptase to form the enzymat ically active tetramer was more dependent on a highly conserved Trp-rich do main on its surface. Although recombinant mMCP-6 and mMCP-7 preferred to fo rm homotypic tetramers, these tryptases readily formed heterotypic tetramer s in vitro. This latter finding indicates that the tetramer structural unit is a novel way the mast cell uses to assemble varied combinations of trypt ases.