Super-Resolution Fluorescence Microscopy Reveals Clustering Behaviour of Chlamydia pneumoniae's Major Outer Membrane Protein.
Super-Resolution Fluorescence Microscopy Reveals Clustering Behaviour of Chlamydia pneumoniae's Major Outer Membrane Protein.
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超分辨率荧光显微镜揭示了肺炎衣原体主要外膜蛋白的聚类行为。
DOI：
10.3390/biology9100344
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发表时间：
2020-10-20
期刊：
影响因子：
4.2
通讯作者：
Watson KA
中科院分区：
文献类型：
--
作者：
Danson AE;McStea A;Wang L;Pollitt AY;Martin-Fernandez ML;Moraes I;Walsh MA;MacIntyre S;Watson KA
衣原体是一种臭名昭著的性传播细菌，在人类呼吸道感染中也扮演着不太为人所知的角色，它进化出一种独特的细胞结构，使其能够在体内生存。覆盖在这种感染细胞表面的是一个强大的网状网络，由许多不同的蛋白质组成，保护细胞免受损害。这项研究专注于该网状结构中最丰富的蛋白质--主要外膜蛋白(MOMP)，并引入了一系列旨在防止网状结构完全形成的突变。突变的影响是通过在每个MOMP中添加一种明亮的荧光染料来可视化的，然后用高分辨率荧光显微镜进行检查，该显微镜能够向我们显示每个单独的细胞及其表面的MOMP。通过统计分析，我们观察到某些突变破坏了MOMP之间的连接，使我们更深入地了解衣原体是如何形成这些相互作用的。衣原体是一种在全球人口中极其流行的疾病，虽然是可以治疗的，但目前还没有可用的疫苗。通过研究衣原体的生物学及其逃避免疫系统的方法，不仅可以加深对这种复杂细菌的了解，还可以为其治疗和预防开发新的治疗方法。肺炎衣原体是一种革兰氏阴性细菌，可导致许多人类呼吸道疾病，并与一些慢性炎症性疾病有关。衣原体主要外膜蛋白(MOMP)是位于外膜的一种保守的免疫优势蛋白，加上其表面暴露和丰富，使其成为近几十年来疫苗和抗菌药物研究的主要焦点。MOMP通过分子间二硫键的形成在衣原体外膜复合体中发挥主要作用，尽管目前尚不清楚形成的确切相互作用。在这里，有人提出，由于大量的半胱氨酸可用于二硫键，相互作用发生在富含半胱氨酸的口袋之间，而不是单个残基。使用具有支持低分辨率(~4？)晶体结构的MOMP同源模型来识别这样的口袋。利用直接随机光学重建显微镜(DSTORM)对MOMP在大肠杆菌膜中的定位进行了评估，结果表明膜上的聚集性减少，富含半胱氨酸的区域包含两个突变。这些结果表明，位于半胱氨酸密集区的单个突变体没有破坏二硫键的形成，而是被口袋中邻近的半胱氨酸补偿，这支持了这一富含半胱氨酸的口袋假说。
Chlamydia is an infamous sexually transmitted bacterium that also has a less well-known role in human respiratory infections, which has evolved a unique cell structure to enable its survival within the body. Covering the surface of this infectious cell is a strong mesh-like network made up of many different proteins which protects the cell against damage. This research focussed on the most abundant protein within this mesh, the Major Outer Membrane Protein (MOMP), and introduced a series of mutations designed to prevent the mesh from forming completely. The effect of the mutations was visualised by adding a bright fluorescent dye to each MOMP, which was then examined with a high-resolution fluorescence microscope capable of showing us each individual cell and the MOMPs at their surface. With statistical analysis, we observed that certain mutations disrupted the connections between MOMPs, giving us greater insight into how Chlamydia forms these interactions. Chlamydia is an extremely prevalent disease amongst the global population, and whilst treatable, there is currently no available vaccine. By researching Chlamydia’s biology and its method of evading our immune system, we can not only further our understanding of this complex bacterium, but also develop novel therapeutics for its treatment and prevention. Chlamydia pneumoniae is a Gram-negative bacterium responsible for a number of human respiratory diseases and linked to some chronic inflammatory diseases. The major outer membrane protein (MOMP) of Chlamydia is a conserved immunologically dominant protein located in the outer membrane, which, together with its surface exposure and abundance, has led to MOMP being the main focus for vaccine and antimicrobial studies in recent decades. MOMP has a major role in the chlamydial outer membrane complex through the formation of intermolecular disulphide bonds, although the exact interactions formed are currently unknown. Here, it is proposed that due to the large number of cysteines available for disulphide bonding, interactions occur between cysteine-rich pockets as opposed to individual residues. Such pockets were identified using a MOMP homology model with a supporting low-resolution (~4 Å) crystal structure. The localisation of MOMP in the E. coli membrane was assessed using direct stochastic optical reconstruction microscopy (dSTORM), which showed a decrease in membrane clustering with cysteine-rich regions containing two mutations. These results indicate that disulphide bond formation was not disrupted by single mutants located in the cysteine-dense regions and was instead compensated by neighbouring cysteines within the pocket in support of this cysteine-rich pocket hypothesis.
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影响因子：
48
作者：
Dempsey, Graham T.;Vaughan, Joshua C.;Chen, Kok Hao;Bates, Mark;Zhuang, Xiaowei
通讯作者：
Zhuang, Xiaowei
影响因子：
3.7
作者：
Feher VA;Randall A;Baldi P;Bush RM;de la Maza LM;Amaro RE
通讯作者：
Amaro RE
影响因子：
5.9
作者：
Hahn, DL;McDonald, R
通讯作者：
McDonald, R
影响因子：
2.2
作者：
Pourhajibagher, M.;Bahador, A.
通讯作者：
Bahador, A.
影响因子：
3.7
作者：
Atanu, Francis O.;Oviedo-Orta, Ernesto;Watson, Kimberly A.
通讯作者：
Watson, Kimberly A.