Mountfort D O, Mörschel E, Beimborn D B, Schönheit P
J Bacteriol. 1986 Nov;168(2):892-900. doi: 10.1128/jb.168.2.892-900.1986.
When Methanobacterium thermoautotrophicum cells were incubated in 50 mM potassium phosphate buffer (pH 7.0) containing 1 M sucrose and autolysate from Methanobacterium wolfei, they were transformed into protoplasts. The protoplasts, which possessed no cell wall, lysed in buffer without sucrose. Unlike whole cells, the protoplasts did not show convoluted internal membrane structures. The protoplasts produced methane from H2-CO2 (approximately 1 mumol min-1 mg of protein-1) at about 50% the rate obtained for whole cells, and methanogenesis was coupled with ATP synthesis. Addition of the protonophore 3,5-di-tert-butyl-4-hydroxybenzylidenemalononitrile (SF-6847) to protoplast suspensions resulted in a dissipation of the membrane potential (delta psi), and this was accompanied by a parallel decrease in the rates of ATP synthesis and methanogenesis. In this respect protoplasts differed from whole cells in which ATP synthesis and methanogenesis were virtually unaffected by the addition of the protonophore. It is concluded that the insensitivity of whole cells to protonophores could be due to internal membrane structures. Membrane preparations produced from lysis of protoplasts or by sonication of whole cells gave comparatively low rates of methanogenesis (methylcoenzyme M methylreductase activity, less than or equal to 100 nmol of CH4 min-1 mg of protein-1), and no coupling with ATP synthesis could be demonstrated.
将嗜热自养甲烷杆菌细胞在含有1 M蔗糖和沃氏甲烷杆菌自溶产物的50 mM磷酸钾缓冲液（pH 7.0）中培养时，它们会转化为原生质体。没有细胞壁的原生质体在不含蔗糖的缓冲液中会裂解。与完整细胞不同，原生质体没有呈现出卷曲的内膜结构。原生质体从H2-CO2产生甲烷（约1 μmol min-1 mg蛋白质-1），其速率约为完整细胞的50%，并且甲烷生成与ATP合成相偶联。向原生质体悬浮液中添加质子载体3,5-二叔丁基-4-羟基苄叉丙二腈（SF-6847）会导致膜电位（Δψ）的消散，同时ATP合成速率和甲烷生成速率也会相应降低。在这方面，原生质体与完整细胞不同，完整细胞中添加质子载体对ATP合成和甲烷生成几乎没有影响。可以得出结论，完整细胞对质子载体不敏感可能是由于内膜结构。由原生质体裂解或通过超声破碎完整细胞制备的膜制剂产生的甲烷生成速率相对较低（甲基辅酶M甲基还原酶活性，≤100 nmol CH4 min-1 mg蛋白质-1），并且无法证明与ATP合成相偶联。
