Structural Stability of Vault Particles
Structural Stability of Vault Particles
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DOI：
10.1002/jps.21508
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发表时间：
2009-04-01
影响因子：
3.8
通讯作者：
Middaugh, C. Russell
中科院分区：
文献类型：
--
作者：
Esfandiary, Reza;Kickhoefer, Valerie A.;Middaugh, C. Russell
穹窿的大小为 13 MDa，是已知最大的核糖核蛋白颗粒。在体外，主要穹窿蛋白 (MVP) 在 Sf9 昆虫细胞中单独表达，导致产生具有特征穹窿结构的重组颗粒。为了利用重组穹窿作为纳米胶囊来递送生物分子，我们采用了各种光谱技术（即圆二色性、荧光光谱和光散射）以及电子显微镜，来表征穹窿在广泛的 pH (3-8) 和温度 (10-90 摄氏度) 范围内的结构稳定性。在所研究的 pH 值和温度范围内，鉴定出拱顶的 10 种不同构象状态，其中在 40 摄氏度以下的 pH 6-8 区域最稳定，在 60 摄氏度以上的 pH 4-6 区域最不稳定。在 pH 6 和类似于 55 摄氏度的情况下还鉴定出一种独特的中间熔球状状态。EM 成像显示，当 pH 值从中性过渡到酸性时，完整的拱顶开放为花状结构。该信息在将重组穹窿开发成用于药物递送的纳米胶囊方面具有潜在用途，因为可以释放包埋在穹窿中的治疗剂的一种机制是通过打开完整的穹窿结构。 (C) 2008 Wiley-Liss, Inc. 和美国药剂师协会 J Pharm Sci 98:1376-1386, 2009
Vaults, at 13 MDa, are the largest ribonucleoprotein particles known. In vitro, expression of the major vault protein (MVP) alone in Sf9 insect cells results in the production of recombinant particles with characteristic vault structure. With the ultimate goal of using recombinant vaults as nanocapsules for the delivery of biomolecules, we have employed a variety of spectroscopic techniques (i.e., circular dichroism, fluorescence spectroscopy, and light scattering) along with electron microscopy, to characterize the structural stability of vaults over a wide range of pH (3-8) and temperature (10-90 degrees C). Ten different conformational states of the vaults were identified over the pH and temperature range studied with the most stable region at pH 6-8 below 40 degrees C and least stable at pH 4-6 above 60 degrees C. A unique intermediate molten globulelike state was also identified at pH 6 and similar to 55 degrees C. EM imaging showed the opening of intact vaults into flowerlike structures when transitioning from neutral to acidic pH. This information has potential use in the development of recombinant vaults into nanocapsules for drug delivery since one mechanism by which therapeutic agents entrapped in vaults could be released is through an opening of the intact vault structure. (C) 2008 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 98:1376-1386, 2009