PURIFICATION AND CHARACTERIZATION OF A NOVEL ISOFORM OF MAST-CELL TRYPTASE FROM RAT TONGUE

Rat mast cell tryptase was purified to homogeneity from rat tongue by a series of standard chromatographic procedures. Since the enzyme gave band corresponding to molecular mass of 32-35 kDa on sodium dodecyl s ulfate polyacrylamide gel electrophoresis and exhibited a molecular ma ss of 135 kDa on gel filtration, it was presumed to be a noncovalently associated tetramer, The N-terminal amino acid sequence of 50 residue s of the enzyme showed the highest degree of homology with the same re gion in mouse mast cell protease 7 (92%), and less homology to those o f tryptases from man and dog, and peritoneal cells of rats and Mongoli an gerbils, The inhibitor specificity of rat tongue tryptase was simil ar to that of rat peritoneal mast cell tryptase free from trypstatin: it was inhibited by alpha(1)-antitrypsin, Kunitz-type soybean trypsin inhibitor and Bowman-Birk soybean trypsin inhibitor, but these inhibit ors do not inhibit the tryptases from rat skin, human lung, and dog ma st cells, Judging from these results, together with other enzymatic pr operties, the enzyme may be a novel isoform of tryptase in rat tongue, Analysis by differential staining with peroxidase-labeled lectins of the enzyme suggested that it has tri- and/or tetraantennary complex-ty pe oligosaccharides containing a relatively high amount of sialic acid , The immunohistochemical distribution of this enzyme indicated that t he reactive antigen was specific in connective tissue but not in mucos al mast cells.