Molecular and functional characterization of channel catfish (Ictalurus punctatus) neutrophil collagenase

Channel catfish (Ictalurus punctatus) neutrophils, like mammalian neutrophi ls, contain a Variety of enzymes and lytic peptides that participate in pat hogen destruction. We have identified and characterized from a channel catf ish anterior kidney cDNA library a 1.6 kb cDNA that encodes for channel cat fish neutrophil collagenase. The deduced amino acid sequence has a predicte d molecular mass of 53 kDa. The putative catfish collagenase has nucleotide and amino acid homology of 51.4% and 45.1%, respectively, with human neutr ophil collagenase and 50.4% and 47.1%, respectively, with mouse neutrophil collagenase. Certain regions of the molecule, including the cysteine switch and the putative zinc binding sites, were identical to those in the human and mouse genes. Polyclonal antiserum, prepared to the fusion protein, reco gnizes proteins from channel catfish neutrophil supernatants with molecular masses of approximately 63, 53 and 28 kDa. Supernatants from phorbol dibut yrate stimulated neutrophils were capable of degrading type I collagen. In addition, the polyclonal antiserum prevented the collagenase activity of th e supernatants from stimulated catfish neutrophils; whereas, preimmune seru m had no effect on collagenase activity of supernatants. Supernatants from unstimulated cells or the fusion protein did not possess the ability of deg rading type I collagen. These results indicate that channel catfish neutrop hil collagenases share molecular and functional features with mammalian neu trophil collagenase. (C) 1999 Elsevier Science B.V. All rights reserved.