Purification and characterization of a new tissue inhibitor of metalloproteinases (TIMP-2) from mouse colon 26 tumor cells.
Mouse colon 26 tumor cells were shown to produce collagenase inhibitor in culture. The inhibitor was purified more than 2,000-fold from the culture medium by passage through DE-52 cellulose, CM-52 cellulose, Ultrogel AcA 54, Con A-Sepharose, and Sephadex G-50 Superfine columns. The inhibitor did not bind to Con A-Sepharose as do most other collagenase inhibitors. The inhibitor showed a single band (Mr = 20.5 k) on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, and inhibitory activity against interstitial collagenases and gelatinases, except for bacterial collagenase. Double-immunodiffusion analysis using monospecific anti-serum against tissue inhibitor of metalloproteinases (TIMP) from bovine dental pulp showed that colon 26 inhibitor did not cross-react immunologically with the pulp inhibitor. NH2-Terminal protein sequence data were obtained for the first 36 residues of the colon 26 inhibitor, and the first 20 of them exhibited a sequence almost identical with that of a new TIMP recently designated as TIMP-2.
Kishi
J
J
Department of Biochemistry, School of Dentistry, Aichi-Gakuin University, Nagoya, Japan.
Ogawa
K
K
Yamamoto
S
S
Hayakawa
T
T
D000230:Adenocarcinoma / Q000503:physiopathology*; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D002846:Chromatography, Affinity; D002850:Chromatography, Gel / Q000379:methods; D002852:Chromatography, Ion Exchange; D003110:Colonic Neoplasms / Q000503:physiopathology*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D018093:Gelatinases; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D003012:Microbial Collagenase / Q000037:antagonists & inhibitors*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009363:Neoplasm Proteins / Q000302:isolation & purification* / Q000494:pharmacology; D010434:Pepsin A / Q000037:antagonists & inhibitors*; D012689:Sequence Homology, Nucleic Acid; D019716:Tissue Inhibitor of Metalloproteinase-2
D009363:Neoplasm Proteins; D019716:Tissue Inhibitor of Metalloproteinase-2; D010434:Pepsin A; D018093:Gelatinases; D003012:Microbial Collagenase