Formation of active monomers from tetrameric human beta-tryptase.
Tryptase is a serine protease that is stored at low pH in the mast cell secretory granules in complex with heparin proteoglycan. When mast cells are activated, e.g. during allergic responses, the tryptase/heparin complexes are released together with a variety of other preformed inflammatory mediators. Previous crystallization of human beta-tryptase revealed a unique tetrameric structure with all of the active sites facing a central pore that has a limited accessibility both for potential substrates as well as for protease inhibitors. In this study we examined whether human beta-tryptase, in addition, could form active monomers. Incubation of recombinant tetrameric human beta-tryptase at neutral pH and 37 degrees C, followed by gel-filtration analysis using a running buffer containing pig mucosal heparin, led to the formation of enzymically active compounds that were of a size compatible with tryptase monomers in complex with heparin. The monomers were, in contrast to tryptase in the tetrameric form, inhibited by bovine pancreatic trypsin inhibitor. Further, the monomers, but not the tetramers, degraded fibronectin. Formation of active monomers was more pronounced at pH 7.5 than at pH 6.0 and was not detected at room temperature or at high heparin/tryptase ratios. The present findings thus introduce the possibility that human beta-tryptase, after mast cell degranulation and exposure to neutral pH in the tissue, may dissociate into active monomers with properties that are distinct from the tetrameric counterpart. Possibly, some of the biological activities of human tryptase may be attributable to active tryptase in its monomeric rather than tetrameric form.
Fajardo
Ignacio
I
Swedish University of Agricultural Sciences, Department of Veterinary Medical Chemistry, The Biomedical Center, Box 575, 751 23 Uppsala, Sweden.
Pejler
Gunnar
G
D007611:Aprotinin / Q000494:pharmacology; D019281:Dimerization; D004795:Enzyme Stability; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008407:Mast Cells / Q000378:metabolism; D008970:Molecular Weight; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D053802:Tryptases
D005353:Fibronectins; D015842:Serine Proteinase Inhibitors; D007611:Aprotinin; D012697:Serine Endopeptidases; D053802:Tryptases