Benz R, Maier E, Ladant D, Ullmann A, Sebo P
Lehrstuhl fur Biotechnologie, Theodor-Boveri-Institut (Biozentrum), Universitat Wurzburg, Federal Republic of Germany.
J Biol Chem. 1994 Nov 4;269(44):27231-9.
The interaction between the adenylate cyclase toxin (CyaA) of Bordetella pertussis and lipid was studied using the lipid bilayer assay. The addition of CyaA to the aqueous phase bathing lipid bilayer membranes composed of different lipids resulted in the increase of the membrane conductance. This increase was rather small for membranes formed of pure lipids as compared with lipid mixtures such as asolectin. The toxin formed in asolectin membranes small transient ion-permeable channels with a single-channel conductance of 27 pS in 1 M KCl, which is considerably smaller than that of the alpha-hemolysin (HlyA) of Escherichia coli (1500 pS). Experiments with different salts suggested that the CyaA-induced channels were exclusively cation-selective because of negative charges localized at the channel mouth. The single-channel conductance of channels initiated by CyaA was independent of whether the toxin was purified from B. pertussis or from recombinant E. coli. However, the channel-forming activity of the CyaA expressed in B. pertussis was substantially higher than that of the recombinant toxin. Experiments with mutant forms of CyaA suggested that both the activation of CyaA by CyaC and the hemeolytic part of the toxin, but not the repeats and the cyclase activity, are required for channel formation in lipid bilayer membranes.
利用脂质双层分析研究了百日咳博德特氏菌腺苷酸环化酶毒素（CyaA）与脂质之间的相互作用。将CyaA添加到浸泡由不同脂质组成的脂质双层膜的水相中，会导致膜电导增加。与诸如大豆卵磷脂等脂质混合物相比，由纯脂质形成的膜的这种增加相当小。毒素在大豆卵磷脂膜中形成了小的瞬时离子通透通道，在1 M KCl中其单通道电导为27 pS，这比大肠杆菌的α-溶血素（HlyA）（1500 pS）小得多。用不同盐进行的实验表明，由于通道口处存在负电荷，CyaA诱导的通道具有阳离子选择性。由CyaA引发的通道的单通道电导与毒素是从百日咳博德特氏菌还是从重组大肠杆菌中纯化出来无关。然而，在百日咳博德特氏菌中表达的CyaA的通道形成活性明显高于重组毒素。对CyaA突变体形式的实验表明，CyaC对CyaA的激活以及毒素的溶血部分，而非重复序列和环化酶活性，是脂质双层膜中通道形成所必需的。
