Identification of a proteolytic activity which responds to anticarcinogenic protease inhibitors in C3H-10T1/2 cells.
Untransformed and malignantly-transformed mouse embryo fibroblasts were found to contain an enzymatic activity which hydrolysed the synthetic substrate, Suc-Ala-Ala-Pro-Phe-AMC. This activity, of approximate molecular weight 55,000, which has been partially purified by ion-exchange and gel-filtration chromatography, was maximally active at neutral pH, associated with subcellular organelles or membranes and inhibited by EDTA, EGTA, phosphoramidon and 1,10 phenanthroline, but not by PMSF or pepstatin, indicating that it may be a metalloprotease. Several other protease inhibitors, such as chymostatin, TPCK and the BBI from soybean, were also potent inhibitors of the activity; these same inhibitors are known to suppress the malignant transformation of mouse embryo fibroblast cells in vitro induced by X-irradiation or chemical carcinogens. In contrast, SBTI and pepstatin, which did not inhibit this activity, also do not suppress malignant transformation. These results suggest that this enzyme may be involved in attainment of the transformed state.
Carew
J A
JA
Harvard School of Public Health, Boston, MA 02115.
Kennedy
A R
AR
D000818:Animals; D002460:Cell Line; D002471:Cell Transformation, Neoplastic / Q000187:drug effects* / Q000378:metabolism; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D005347:Fibroblasts / Q000201:enzymology; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008666:Metalloendopeptidases / Q000378:metabolism*; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology*; D013347:Subcellular Fractions / Q000201:enzymology; D013379:Substrate Specificity
D011480:Protease Inhibitors; D008666:Metalloendopeptidases; C063182:succinylalanyl-alanyl-prolyl-phenylalanyl-7-amino-4-methylcoumarin hydrolase