Krag S S, Lennarz W J
J Biol Chem. 1975 Apr 25;250(8):2813-22.
The protoplasts of a mutant of Bacillus subtilis 168 (B. subtilis CMK33) are osmotically fragile when compared to protoplasts of the parent organism and contain an active, membrane-associated phospholipase A1. A protein found in the parent organism specifically inhibits the phospholipase A1 (Kent, C., and Lennarz, W.J. (1972) Proc. Nat. Acad. Sci. U.S.A. 69, 2793-2797). The inhibitor exists in both a soluble and particulate form. The soluble inhibitor is not found in the cytoplasm, but rather in a "periplasmic" fraction released from the cell during incubation with lysozyme. The soluble inhibitor has been purified to homogeneity by diethylaminoethyl-cellulose and hydroxylapatite chromatography. Its molecular weight is 28,000 to 32,000 as determined by gel filtration chromatography and 36,000 to 37,000 as determined by sodium dodecyl sulfate-urea gel electrophoresis. The inhibitor appears to inactivate the membrane bound phospholipase A1 by an enzymatic process that is dependent on time and protein concentration. Binding of the inhbitor to the membrane-associated phospholipase cannot be detected. When purified inhibitor is added to cells of B. subtilis CMK33 during treatment with lysozyme, the osmotic stability of the resultant protoplasts is similar to that of protoplasts of the wild type of organism.
与亲本菌株的原生质体相比，枯草芽孢杆菌168的一个突变体（枯草芽孢杆菌CMK33）的原生质体对渗透压敏感，并且含有一种活性的、与膜相关的磷脂酶A1。在亲本菌株中发现的一种蛋白质可特异性抑制磷脂酶A1（肯特，C.，和伦纳兹，W.J.（1972年）《美国国家科学院院刊》69，2793 - 2797）。该抑制剂以可溶性和颗粒性两种形式存在。可溶性抑制剂不存在于细胞质中，而是存在于与溶菌酶孵育期间从细胞中释放出的“周质”部分。可溶性抑制剂已通过二乙氨基乙基纤维素和羟基磷灰石层析纯化至同质。通过凝胶过滤层析测定其分子量为28,000至32,000，通过十二烷基硫酸钠 - 尿素凝胶电泳测定为36,000至37,000。该抑制剂似乎通过一个依赖于时间和蛋白质浓度的酶促过程使膜结合的磷脂酶A1失活。未检测到抑制剂与膜相关磷脂酶的结合。当在溶菌酶处理期间将纯化的抑制剂添加到枯草芽孢杆菌CMK33的细胞中时，所得原生质体的渗透稳定性与野生型菌株原生质体的相似。
