Isolation and characterisation of a chicken gelatinase (type IV collagenase).
The proform of chick gelatinase (type IV collagenase) was isolated and purified to a high specific activity of 12,071 U/mg from cultured embryonic skin fibroblasts stimulated with cytochalasin-B. The enzyme was activated in the presence of 4-aminophenylmercuric acetate with a fall in molecular weight from 66,000-58,000 on non-reducing polyacrylamide gel electrophoresis and was active over the pH range of 6.0-8.9 against a number of substrates. Further biochemical characterisation showed that the organomercurial activated form of the enzyme behaved like a typical mammalian gelatinase, actively degrading gelatin, soluble type I collagen, collagenase generated type I fragments, type IV collagen (producing 3/4 and 1/4 fragments) and type V collagen, whilst having little effect on laminin. The enzyme was inhibited by metal chelators such as EDTA and 1,10-phenanthroline, but not by inhibitors is suggested that this may be TIMP-2. An antiserum was raised to the proenzyme and was found to localise intra- and extra-cellularly in both tissue sections and cell cultures.
Craig
F M
FM
University Department of Orthopaedic Surgery, University College and Middlesex Schools of Medicine, RNOH, Stanmore, U.K.
Archer
C W
CW
Murphy
G
G
D000818:Animals; D002642:Chick Embryo / Q000737:chemistry* / Q000187:drug effects; D003094:Collagen / Q000378:metabolism; D003571:Cytochalasin B / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005780:Gelatin / Q000378:metabolism; D006023:Glycoproteins / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007797:Laminin / Q000378:metabolism; D020780:Matrix Metalloproteinase 9; D003012:Microbial Collagenase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D012867:Skin / Q000737:chemistry* / Q000187:drug effects / Q000196:embryology; D019714:Tissue Inhibitor of Metalloproteinases
D006023:Glycoproteins; D007797:Laminin; D019714:Tissue Inhibitor of Metalloproteinases; D003571:Cytochalasin B; D005780:Gelatin; D003094:Collagen; D003012:Microbial Collagenase; D020780:Matrix Metalloproteinase 9