Smith D K, Winkler H H
J Bacteriol. 1979 Feb;137(2):963-71. doi: 10.1128/jb.137.2.963-971.1979.
Rickettsia prowazeki were disrupted in a French pressure cell and fractionated into soluble (cytoplasm) and envelope fractions. The envelope contained 25% of the cell protein, with the cytoplasm containing 75%. Upon density gradient centrifugation, the envelope fraction separated into a heavy band (1.23 g/cm3) and a lighter band (1.19 g/cm3). The heavy band had a high content of 2-keto-3-deoxyoctulosonic acid, a marker for bacterial lipopolysaccharide, but had no succinic dehydrogenase, a marker for cytoplasmic membrane activity, and therefore represented outer membrane. The lighter band exhibited a high succinate dehydrogenase activity, and thus contained inner (cytoplasmic) membrane. Outer membrane purified by this method was less than 5% contaiminated by cytoplasmic membrane; however, inner membrane from the gradient was as much as 30% contaminated by outer membrane. The protein composition of each cellular fraction was characterized by sodium dodecyl sulfate--polyacrylamide gel electrophoresis. The outer membrane contained four major proteins, which were also major proteins of the whole cell. The cytoplasmic membrane and soluble cytoplasm exhibited a more complex pattern on gels.
普氏立克次体在法国压力室中被破碎，并分离成可溶性（细胞质）和包膜组分。包膜含有25%的细胞蛋白，细胞质含有75%。在密度梯度离心时，包膜组分分离成一条重带（1.23 g/cm³）和一条较轻的带（1.19 g/cm³）。重带含有高含量的2-酮-3-脱氧辛糖酸，这是细菌脂多糖的标志物，但没有琥珀酸脱氢酶，这是细胞质膜活性的标志物，因此代表外膜。较轻的带表现出高琥珀酸脱氢酶活性，因此含有内膜（细胞质膜）。用这种方法纯化的外膜被细胞质膜污染不到5%；然而，梯度中的内膜被外膜污染高达30%。每个细胞组分的蛋白质组成通过十二烷基硫酸钠-聚丙烯酰胺凝胶电泳进行表征。外膜含有四种主要蛋白质，它们也是全细胞的主要蛋白质。细胞质膜和可溶性细胞质在凝胶上呈现出更复杂的图谱。
