Kishi J, Ogawa K, Yamamoto S, Hayakawa T
Department of Biochemistry, School of Dentistry, Aichi-Gakuin University, Nagoya, Japan.
Matrix. 1991 Feb;11(1):10-6. doi: 10.1016/s0934-8832(11)80222-2.
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.
小鼠结肠26肿瘤细胞在培养过程中可产生胶原酶抑制剂。通过DE - 52纤维素柱、CM - 52纤维素柱、Ultrogel AcA 54柱、伴刀豆球蛋白A - 琼脂糖柱和Sephadex G - 50超细柱从培养基中对该抑制剂进行了2000多倍的纯化。该抑制剂不像大多数其他胶原酶抑制剂那样与伴刀豆球蛋白A - 琼脂糖结合。在十二烷基硫酸钠（SDS）-聚丙烯酰胺凝胶电泳上，该抑制剂呈现单一条带（相对分子质量=20.5k），且除细菌胶原酶外，对间质胶原酶和明胶酶具有抑制活性。使用针对牛牙髓金属蛋白酶组织抑制剂（TIMP）的单特异性抗血清进行双向免疫扩散分析表明，结肠26抑制剂与牙髓抑制剂无免疫交叉反应。获得了结肠26抑制剂前36个残基的氨基末端蛋白质序列数据，其中前20个残基的序列与最近命名为TIMP - 2的一种新TIMP的序列几乎相同。
