Kusunoki Hideki, Tanaka Toshiyuki, Kohno Toshiyuki, Kimura Hirokazu, Hosoda Kazuo, Wakamatsu Kaori, Hamaguchi Isao
Department of Research on Blood and Biological Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan.
Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan.
Biochem Biophys Rep. 2016 Dec 24;9:159-165. doi: 10.1016/j.bbrep.2016.12.006. eCollection 2017 Mar.
Hepatitis B virus X protein (HBx) is a multifunctional protein that interacts directly with many host proteins. For example, HBx interacts with anti-apoptotic proteins, Bcl-2 and Bcl-x, through its BH3-like motif, which leads to elevated cytosolic calcium levels, efficient viral DNA replication and the induction of apoptosis. To facilitate sample preparation and perform detailed structural characterization of the complex between HBx and Bcl-x, we designed and purified a recombinant HBx BH3-like motif-linker-Bcl-x fusion protein produced in . The fusion protein was characterized by size exclusion chromatography, circular dichroism and nuclear magnetic resonance experiments. Our results show that the fusion protein is a monomer in aqueous solution, forms a stable intramolecular complex, and likely retains the native conformation of the complex between Bcl-x and the HBx BH3-like motif. Furthermore, the HBx BH3-like motif of the intramolecular complex forms an α-helix. These observations indicate that the fusion protein should facilitate structural studies aimed at understanding the interaction between HBx and Bcl-x at the atomic level.
乙型肝炎病毒X蛋白（HBx）是一种多功能蛋白，可直接与许多宿主蛋白相互作用。例如，HBx通过其BH3样基序与抗凋亡蛋白Bcl-2和Bcl-x相互作用，这会导致胞质钙水平升高、有效的病毒DNA复制以及凋亡的诱导。为了便于样品制备并对HBx与Bcl-x之间的复合物进行详细的结构表征，我们设计并纯化了一种在……中产生的重组HBx BH3样基序-连接子-Bcl-x融合蛋白。该融合蛋白通过尺寸排阻色谱、圆二色性和核磁共振实验进行表征。我们的结果表明，该融合蛋白在水溶液中是单体，形成稳定的分子内复合物，并且可能保留了Bcl-x与HBx BH3样基序之间复合物的天然构象。此外，分子内复合物的HBx BH3样基序形成一个α螺旋。这些观察结果表明，该融合蛋白应有助于旨在在原子水平上理解HBx与Bcl-x之间相互作用的结构研究。
