Sagane Yoshimasa, Mutoh Shingo, Koizumi Ryosuke, Suzuki Tomonori, Miyashita Shin-Ichiro, Miyata Keita, Ohyama Tohru, Niwa Koichi, Watanabe Toshihiro
Department of Food and Cosmetic Science, Faculty of Bioindustry, Tokyo University of Agriculture, 196 Yasaka, Abashiri, 099-2493, Japan.
Department of Health and Nutrition, Faculty of Human Science, Hokkaido Bunkyo University, 5-196-1 Kogane-chuo, Eniwa, 061-1449, Japan.
Protein J. 2017 Oct;36(5):417-424. doi: 10.1007/s10930-017-9733-y.
Botulinum neurotoxin (BoNT) associates with nontoxic proteins, either a nontoxic nonhemagglutinin (NTNHA) or the complex of NTNHA and hemagglutinin (HA), to form M- or L-toxin complexes (TCs). Single BoNT and NTNHA molecules are associated and form M-TC. A trimer of the 70-kDa HA protein (HA-70) attaches to the M-TC to form M-TC/HA-70. Further, 1-3 arm-like 33- and 17-kDa HA molecules (HA-33/HA-17 trimer), consisting of 1 HA-17 protein and 2 HA-33 proteins, can attach to the M-TC/HA-70 complex, yielding 1-, 2-, and 3-arm L-TC. In this study, the purified 1- and 2-arm L-TCs spontaneously converted into another L-TC species after acquiring the HA-33/HA-17 trimer from other TCs during long-term storage and freezing/thawing. Transmission electron microscopy analysis provided evidence of the formation of detached HA-33/HA-17 trimers in the purified TC preparation. These findings provide evidence of reversible association/dissociation of the M-TC/HA-70 complex with the HA-33/HA-17 trimers, as well as dynamic conversion of the quaternary structure of botulinum TC in culture.
肉毒杆菌神经毒素（BoNT）与无毒蛋白结合，该无毒蛋白可以是无毒非血凝素（NTNHA），也可以是NTNHA与血凝素（HA）的复合物，从而形成M型或L型毒素复合物（TCs）。单个BoNT分子与NTNHA分子结合形成M型毒素复合物。70 kDa HA蛋白（HA-70）的三聚体附着在M型毒素复合物上形成M型毒素复合物/HA-70。此外，由1个HA-17蛋白和2个HA-33蛋白组成的1-3个臂状33 kDa和17 kDa HA分子（HA-33/HA-17三聚体）可以附着在M型毒素复合物/HA-70复合物上，产生1臂、2臂和3臂L型毒素复合物。在本研究中，纯化的1臂和2臂L型毒素复合物在长期储存以及冻融过程中从其他毒素复合物获得HA-33/HA-17三聚体后，会自发转化为另一种L型毒素复合物。透射电子显微镜分析提供了纯化的毒素复合物制剂中游离HA-33/HA-17三聚体形成的证据。这些发现提供了M型毒素复合物/HA-70复合物与HA-33/HA-17三聚体可逆结合/解离的证据，以及肉毒杆菌毒素复合物在培养过程中四级结构动态转化的证据。
