Purification and N-terminal amino acid sequence of sheep neutrophil cathepsin G and elastase

Sheep cathepsin G (CG) and neutrophil elastase (NE) were isolated from a cr ude leukocyte membrane preparation by elastin-Sepharose 4B and CM:Sepharose 4B chromatography, followed by native preparative PAGE. The N-termini of C G; and NE were sequenced to 24 and 20 residues, showing 96 and 85% identity with human CG and NE, respectively. During SDS-PAGE, sheep CG and NE migra ted parallel to human CG and NE and have apparent molecular masses of 28 an d 26 kDa, respectively. Following incubation of sheep CG and NE with human alpha(1)-antichymotrypsin and alpha(1)-proteinase inhibitor, complexes with apparent molecular masses of 89 and 81 kDa respectively were observed by S DS-PAGE, Polyclonal antibodies to human CG and NE cross-reacted with purifi ed sheep CG and NE, respectively. These results indicate that sheep neutrop hils contain CG and elastase that are analogous to human CG and NE in terms of molecular mass, reactivity with endogenous inhibitors, immunocross-reac tivity, and N-terminal sequence. (C) 1999 Academic Press.