Characterisation of tryptase and a granzyme H-like chymase isolated from equine mastocytoma tissue.
Mast cell proteinases are important inflammatory mediators in man and other species, but until now there has been no investigation of the nature of equine mast cell proteinases. These studies describe the purification and characterisation of two proteolytic components from equine mastocytoma tissue, detected using chromogenic substrates for trypsin and chymotrypsin. Following chromatographic purification, the trypsin-like component was found to be equine mast cell tryptase by N-terminal amino acid sequencing, showing a close similarity with human tryptase-beta (85% identity over 20 residues). It also had similar subunit molecular size (34-36kDa by SDS-PAGE) and substantially similar cleavage specificity to human tryptase-beta with the substrates tested. A 32kDa chymotrypsin-like component was also purified from mastocytoma extract, and termed equine mast cell proteinase-1 (eqMCP-1). The N-terminal amino acid sequence of eqMCP-1 was very similar to human granzyme H (95% over 19 residues). Rabbit antisera directed against tryptase and eqMCP-1 both detected equine mast cells by immunohistochemistry, and will be of use in future clinical studies of the relevance of mast cell proteinases in equine allergic disease.
Pemberton
A D
AD
Department of Veterinary Clinical Studies, Wellcome Trust Centre for Research in Comparative Respiratory Medicine, University of Edinburgh, Easter Bush Veterinary Centre, Roslin, Midlothian EH25 9RG, UK. alan.pemberton@ed.ac.uk
McEuen
A R
AR
Scudamore
C L
CL
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D002846:Chromatography, Affinity / Q000662:veterinary; D002850:Chromatography, Gel / Q000662:veterinary; D053818:Chymases; D006734:Horse Diseases / Q000201:enzymology* / Q000276:immunology; D006736:Horses; D012515:Mast-Cell Sarcoma / Q000201:enzymology / Q000276:immunology / Q000662:veterinary*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011817:Rabbits; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D053802:Tryptases
D012697:Serine Endopeptidases; D053818:Chymases; D053802:Tryptases