Kuwahara Yohta, Unzai Satoru, Nagata Takashi, Hiroaki Yoko, Yokoyama Hideshi, Matsui Ikuo, Ikegami Takahisa, Fujiyoshi Yoshinori, Hiroaki Hidekazu
Field of Supramolecular Biology, International Graduate School of Arts and Sciences, Yokohama City University, Kanagawa, Japan.
Biophys J. 2009 Oct 7;97(7):2034-43. doi: 10.1016/j.bpj.2009.07.034.
Stomatin, prohibitin, flotillin, and HflK/C (SPFH) domain proteins are membrane proteins that are widely conserved from bacteria to mammals. The molecular functions of these proteins have not been established. In mammals, the domain is often found in raft-associated proteins such as flotillin and podocin. We determined the structure of the SPFH domain of PH0470 derived from Pyrococcus horikoshii using NMR. The structure closely resembles that of the SPFH domain of the paralog PH1511, except for two C-terminal helices. The results show that the SPFH domain forms stable dimers, trimers, tetramers, and multimers, although it lacks the coiled-coil region for oligomerization, which is a highly conserved region in this protein family. The oligomers exhibited unusual thermodynamic behavior, as determined by circular dichroism, NMR, gel filtration, chemical cross-linking, and analytical ultracentrifugation. The oligomers were converted into monomers when they were heated once and then cooled. This transition was one-way and irreversible. We propose a mechanism of domain swapping for forming dimers as well as successive oligomers. The results of this study provide what to our knowledge are new insights into the common molecular function of the SPFH domain, which may act as a membrane skeleton through oligomerization by domain swapping.
stomatin、prohibitin、flotillin和HflK/C（SPFH）结构域蛋白是从细菌到哺乳动物广泛保守的膜蛋白。这些蛋白的分子功能尚未明确。在哺乳动物中，该结构域常存在于与脂筏相关的蛋白中，如flotillin和足突蛋白。我们利用核磁共振（NMR）确定了源自嗜热栖热菌的PH0470的SPFH结构域的结构。除了两个C端螺旋外，该结构与旁系同源物PH1511的SPFH结构域非常相似。结果表明，尽管SPFH结构域缺乏该蛋白家族中高度保守的用于寡聚化的卷曲螺旋区域，但它能形成稳定的二聚体、三聚体、四聚体和多聚体。通过圆二色性、核磁共振、凝胶过滤、化学交联和分析超速离心确定，这些寡聚体表现出异常的热力学行为。这些寡聚体一旦加热然后冷却就会转变为单体。这种转变是单向且不可逆的。我们提出了一种结构域交换机制来形成二聚体以及连续的寡聚体。据我们所知，本研究结果为SPFH结构域的共同分子功能提供了新的见解，该结构域可能通过结构域交换进行寡聚化，从而充当膜骨架。
