Partial purification and characterization of exudate gelatinase in the acute phase of carrageenin-induced inflammation in rats.
Gelatinase has been partially purified from exudate in the acute phase of carrageenin-induced inflammation in rats. The enzyme occurs in a latent form that can be activated with 4-aminophenylmercuric acetate (APMA). The latent gelatinase was separated into an active gelatinase and a protein fraction by zinc-chelating Sepharose 6B column chromatography in the final step of purification, suggesting that the latent gelatinase is an enzyme-inhibitor complex. The pH optimum of the active gelatinase is about 7.5 and no reactivity toward native type I collagen or alpha-casein was detected. The molecular weights of the latent and active gelatinases were about 245,000 and about 185,000, respectively, as determined by gel filtration on Sephadex G-200. On the other hand, both latent and active gelatinases occurred in multiple forms in SDS-substrate polyacrylamide gel electrophoresis; the latent gelatinase showed two bands with molecular weights of 105,000 and 69,000, and two additional bands of 88,000 and 83,000 appeared when the latent gelatinase was activated with APMA, while the active gelatinase showed all four species. The active gelatinase was inhibited by metallo-proteinase inhibitors, but not by serine- or cysteine-proteinase inhibitors, suggesting that the exudate gelatinase is a metallo-proteinase. The active gelatinase was also inhibited by serum proteins such as albumin and gamma-globulin, suggesting that gelatinase does not remain in an active form in the inflammatory lesion, where the vascular permeability is increased.
D000210:Acute-Phase Reaction / Q000139:chemically induced / Q000201:enzymology*; D000818:Animals; D002351:Carrageenan; D002845:Chromatography / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D005122:Exudates and Transudates / Q000201:enzymology*; D018093:Gelatinases; D007249:Inflammation / Q000201:enzymology*; D008297:Male; D010434:Pepsin A / Q000302:isolation & purification*; D051381:Rats; D011919:Rats, Inbred Strains; D013379:Substrate Specificity
D002351:Carrageenan; D010434:Pepsin A; D018093:Gelatinases