Sulfolobus acidocaldarius Microvesicles Exhibit Unusually Tight Packing Properties as Revealed by Optical Spectroscopy
Sulfolobus acidocaldarius Microvesicles Exhibit Unusually Tight Packing Properties as Revealed by Optical Spectroscopy
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DOI：
10.3390/ijms20215308
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发表时间：
2019-11-01
影响因子：
5.6
通讯作者：
Chong, Parkson Lee-Gau
中科院分区：
文献类型：
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作者：
Bonanno, Alexander;Blake, Robert C.;Chong, Parkson Lee-Gau
在这项研究中，我们使用光学光谱来表征从嗜热嗜酸古细菌Sulfolobus acidocaldarius(Sa-MVS)释放的微囊的物理性质。Sa-MVS中最丰富的蛋白质是S层蛋白质，它自组装在囊泡表面形成一系列晶体结构。Sa-MVS中的脂类完全是双极四醚类。我们发现，当激发波长为275 nm时，Sa-MVS的本征蛋白质荧光在23℃时的最大发射波长为303 nm(或在75℃时测得的296 nm)，这对于含有多种色氨酸和酪氨酸的蛋白质样品来说是异常低的。在10-11 mM的表面活性剂n-十四烷基-β-d-麦芽糖苷(TDM)存在下，Sa-MVS解体，内源蛋白荧光发射峰向312 nm移动，激发峰由288 nm移至280.5 nm，激发带锐度显著降低(约2倍)。这些数据表明，天然SA-MVS中的大部分荧光氨基酸残基以紧密堆积的蛋白质基质形式存在，S层蛋白质可能形成J-聚集体。用6-十二酰-2-二甲氨基萘(Laurdan)探针研究了Sa-MVS的膜，以及由酸性链球菌极性脂组分E(PLFE)四醚脂(LUVPLFE)、由Sa-MVS提取的四醚脂重组的LUVV(LUVMV)和由双酯脂组成的LUV构成的单层囊泡(LUV)的膜。在紧密堆积的Sa-MVS、LUVMV和LUVPLFE中，Laurdan的广义极化(Gp)值远低于不紧密堆积的DPPC凝胶状态下的Gp，这与先前的发现相一致，即由于探针配置的不同，四醚类脂膜的GP值不能与双酯类脂膜的GP值直接比较。在Sa-MVS和LUVMV中，Laurdan的GP和红边激发位移(REES)值随温度的升高而单调下降，没有脂质相变的迹象。在四醚类脂膜体系(Sa-MVS、LUVMV和LUVPLFE)中，Laurdan‘s Rees值较高(9.3~18.9 nm)，而在双酯脂质体中，Laurdan’s Rees值较低(0.4~5.0 nm)。高的REE和低的GP值表明，Laurdan在四醚类脂膜中，特别是在Sa-MVS膜中，处于一个非常受运动限制的环境中，结合水分子和四醚类脂头基团中的极性部分与Laurdan的激发态偶极矩强烈相互作用，与Laurdan寿命相比，Laurdan在四醚类脂膜中围绕Laurdan发色团的“溶剂”重定向发生得非常慢。
In this study, we used optical spectroscopy to characterize the physical properties of microvesicles released from the thermoacidophilic archaeon Sulfolobus acidocaldarius (Sa-MVs). The most abundant proteins in Sa-MVs are the S-layer proteins, which self-assemble on the vesicle surface forming an array of crystalline structures. Lipids in Sa-MVs are exclusively bipolar tetraethers. We found that when excited at 275 nm, intrinsic protein fluorescence of Sa-MVs at 23 degrees C has an emission maximum at 303 nm (or 296 nm measured at 75 degrees C), which is unusually low for protein samples containing multiple tryptophans and tyrosines. In the presence of 10-11 mM of the surfactant n-tetradecyl-beta-d-maltoside (TDM), Sa-MVs were disintegrated, the emission maximum of intrinsic protein fluorescence was shifted to 312 nm, and the excitation maximum was changed from 288 nm to 280.5 nm, in conjunction with a significant decrease (>2 times) in excitation band sharpness. These data suggest that most of the fluorescent amino acid residues in native Sa-MVs are in a tightly packed protein matrix and that the S-layer proteins may form J-aggregates. The membranes in Sa-MVs, as well as those of unilamellar vesicles (LUVs) made of the polar lipid fraction E (PLFE) tetraether lipids isolated from S. acidocaldarius (LUVPLFE), LUVs reconstituted from the tetraether lipids extracted from Sa-MVs (LUVMV) and LUVs made of the diester lipids, were investigated using the probe 6-dodecanoyl-2-dimethylaminonaphthalene (Laurdan). The generalized polarization (GP) values of Laurdan in tightly packed Sa-MVs, LUVMV, and LUVPLFE were found to be much lower than those obtained from less tightly packed DPPC gel state, which echoes the previous finding that the GP values from tetraether lipid membranes cannot be directly compared with the GP values from diester lipid membranes, due to differences in probe disposition. Laurdan's GP and red-edge excitation shift (REES) values in Sa-MVs and LUVMV decrease with increasing temperature monotonically with no sign for lipid phase transition. Laurdan's REES values are high (9.3-18.9 nm) in the tetraether lipid membrane systems (i.e., Sa-MVs, LUVMV and LUVPLFE) and low (0.4-5.0 nm) in diester liposomes. The high REES and low GP values suggest that Laurdan in tetraether lipid membranes, especially in the membrane of Sa-MVs, is in a very motionally restricted environment, bound water molecules and the polar moieties in the tetraether lipid headgroups strongly interact with Laurdan's excited state dipole moment, and "solvent" reorientation around Laurdan's chromophore in tetraether lipid membranes occurs very slowly compared to Laurdan's lifetime.