Nelson W J, Traub P
Eur J Biochem. 1981 May;116(1):51-7. doi: 10.1111/j.1432-1033.1981.tb05299.x.
A Ca2+-activated neutral protease is described which, when tested against various native proteins, appears to be specific for vimentin, the 58,000-Mr subunit protein of intermediate-sized (7--11 nm) filaments in Ehrlich-ascites-tumour cells. The protein subunits of other classes of intermediate-sized filaments have been tested; neurofilament protein and glial fibrillary acidic protein are not degraded, however skeletin, the subunit protein of intermediate-sized filaments in smooth muscle, is degraded. The protease is found associated with the detergent-resistant cytoskeleton of Ehrlich-ascites-tumour cells; proteins, other than vimentin, present in this structure are not degraded. The protease is activated by Ca2+ and Sr2+ but not by other divalent cations tested: the Ca2+ concentration required for activation is 10 microM. The pH optimum is between pH 7.5 and 8.0, and the KCl concentration required for optimal activity is 100 mM. The protease is inhibited by 1-chloro-3-tosylamido-7-amino-L-2-heptanone hydrochloride and L-1-tosylamido-2-phenylethyl chloromethyl ketone but not by soybean trypsin inhibitor; inhibition by phenylmethylsulphonyl fluoride is moderate. The high substrate specificity of the protease suggests it may play a role in vimentin intermediate-sized filament protein turnover in Ehrlich-ascites-tumour cells.
描述了一种Ca2+激活的中性蛋白酶，当用各种天然蛋白质进行测试时，它似乎对波形蛋白具有特异性，波形蛋白是艾氏腹水瘤细胞中中等大小（7-11纳米）细丝的58,000道尔顿亚基蛋白。已经测试了其他类别的中等大小细丝的蛋白质亚基；神经丝蛋白和胶质纤维酸性蛋白未被降解，然而，平滑肌中中等大小细丝的亚基蛋白肌动蛋白骨架蛋白被降解。发现该蛋白酶与艾氏腹水瘤细胞的耐去污剂细胞骨架相关；存在于该结构中的除波形蛋白以外的蛋白质未被降解。该蛋白酶被Ca2+和Sr2+激活，但不被测试的其他二价阳离子激活：激活所需的Ca2+浓度为10 microM。最适pH在7.5至8.0之间，最佳活性所需的KCl浓度为100 mM。该蛋白酶被盐酸1-氯-3-甲苯磺酰胺基-7-氨基-L-2-庚酮和L-1-甲苯磺酰胺基-2-苯乙基氯甲基酮抑制，但不被大豆胰蛋白酶抑制剂抑制；苯甲基磺酰氟的抑制作用中等。该蛋白酶的高底物特异性表明它可能在艾氏腹水瘤细胞中波形蛋白中等大小细丝蛋白的周转中起作用。
