Mast cells play a central role in the pathogenesis of the allergic response and neutral proteases are the major protein components on a weight basis of the secretory granules of human mast cells. These enzymes are selectively concentrate in mast cells, are released during mast cell degranulation and are the principal focus of this proposed renewal. Tryptase, a serine esterase with trypsin-like substrate specificity, is abundantly present in all mast cells (MCT and MCTC types).
Specific aims concerning human tryptase are four-fold. First, a more sensitive and clinically useful immunoassay for tryptase will be developed as a precise but somewhat insensitive indication of mast cell involvement in natural and experimentally-induced anaphylaxis, asthma, rhinitis and cutaneous allergy. Second, an immunoaffinity purification technique for tryptase will be developed. This will require the production of monoclonal antibodies that recognize the catalytic form of the enzyme and will facilitate purification of substantial quantities of the enzyme to be used to elucidate the structural and functional properties of tryptase. Third, the biologic and physicochemical factors involved in the regulation of tryptase stability, activity and function will be examined. The active from of tryptase is tetrameric, and is stabilized as such by both heparin and chondroitin sulfate E. Without such stabilizers loss of enzymatic activity occurs concomitant with a change in the quaternary structure to a monomeric form. Corresponding changes in secondary structure, for example, will be examined by circular dichroic spectroscopy. The effect of eosinophil basic proteins on tryptase stability and the functional role of tryptase in connective tissue metabolism each will be pursued. Fourth, the number of genes coding for tryptase, existence of common polymorphisms, organization of the tryptase genome and presence of transcriptional regulatory elements will be addressed. Chymase is a serine endopeptidase with specificity for chymotrypsin-like substrates, is monomeric, and resides in the secretory granules of a subset of human mast cells termed MCTC.
Specific aims concerning chymase include cloning and characterization of the gene for chymase in order to determine its genomic organization and identify factors affecting its expression; production of an immunoassay with adequate sensitivity and specificity to measure chymase in biological fluids and thereby indicate involvement of the MCTC type of mast cell; and development of an immunoaffinity purification method. the anticipated accomplishments of this proposal will yield a better understanding of the biology and pathobiology of human mast cells, provide a more precise diagnostic capability concerning allergic diseases and assist with the eventual treatment of such conditions.
| Vegh, Arthur B; George, Kimberly C; Lotfi-Emran, Sahar et al. (2011) Total tryptase levels indicate risk for systemic reactions to rush immunotherapy and mast cell activation. Ann Allergy Asthma Immunol 106:342-343.e6 |
| Simon, Michael R; Jan, Mindy; Yee, Jerry et al. (2010) Tryptase is not cleared by the kidneys into the urine. Int Arch Allergy Immunol 152:28-31 |
