Development of controlling technology for protein dissociation and aggregation using high pressure.

蛋白质聚集是影响人类健康和重组蛋白生产的重要问题之一。本研究的目的是开发利用高压控制蛋白质解离和聚集的技术。在这项研究中，我们做了以下四个主题。1. 利用高压核磁共振波谱技术观察了母鸡溶菌酶的变性过程，在这个过程中，高压抑制了蛋白质的聚集。我们成功地观察到了低洼激发态和平衡状态下展开的中间体（Kamatari et al., Biophys Chem. 156, 24- 30,2011）。2. 我们制作了一个大容量高压装置，并对包涵体进行了折叠式实验。3. 我们制作了高压荧光光谱仪，观察了淀粉样蛋白原纤维的解离过程。4. 除了高压外，我们还使用了化合物来抑制聚合。我们发现了几种新的抗朊病毒化合物(Hosokawa-Muto et al., Antimicrob；代理Chemother。， 53, 765-771, 2009)。我们优化了一种抗朊病毒化合物GN8，发现比GN8效率更高的化合物（Kimura et al., Bioorg Med Chem Lett. 21, 1502-1507, 2011）。

Protein aggregation is one of the serious problems for human health and production of recombinant proteins. Purpose of this study is development of controlling technology for protein dissociation and aggregation using high pressure. In this study, we did following four topics. 1. We observed denaturation process of hen lysozyme using high pressure NMR spectroscopy, in which high pressure suppress protein aggregation. We succeed to observe low-lying excited states and unfolding intermediates at equilibrium condition(Kamatari et al., Biophys Chem. 156, 24-30, 2011). 2. We made a high-volume high pressure apparatus and did refolding experiments from inclusion bodies. 3. We made a high pressure apparatus for fluorescence spectroscopy and observed dissociation process of amyloid fibrils. 4. We used chemical compounds to suppress aggregation in addition to high pressure. We found several new anti-prion compounds(Hosokawa-Muto et al., Antimicrob. Agents Chemother., 53, 765-771, 2009). We optimized a anti-prion compound, GN8, and found higher efficiency compound than GN8(Kimura et al., Bioorg Med Chem Lett. 21, 1502-1507, 2011).