Engineered oligomerization state of OmpF protein through computational design decouples oligomer dissociation from unfolding.
Engineered oligomerization state of OmpF protein through computational design decouples oligomer dissociation from unfolding.
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DOI：
10.1016/j.jmb.2012.02.043
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发表时间：
2012-05-25
影响因子：
5.6
通讯作者：
Liang, Jie
中科院分区：
文献类型：
--
作者：
Naveed, Hammad;Jimenez-Morales, David;Tian, Jun;Pasupuleti, Volga;Kenney, Linda J.;Liang, Jie
关键词：
β-桶膜蛋白的生物发生是一个复杂的、多步骤的、并且尚未完全表征的过程。细菌孔蛋白家族可能是β-桶膜蛋白中研究得最好的蛋白质家族，其允许小溶质扩散穿过细菌外膜。在这项研究中，我们已经确定了残基，有助于显着的蛋白质-蛋白质相互作用（PPI）的外膜蛋白F（OmpF），三聚体孔蛋白的链之间的接口，使用经验能量函数结合进化分析。通过定点诱变替换这些残基，无论是积极有利的残基或取代，不发生在天然细菌外膜蛋白，我们成功地在工程OmpF突变体与二聚体和单体，而不是三聚体寡聚化状态。此外，我们的研究结果表明，OmpF的寡聚化通过一系列的相互作用进行，涉及两个不同的区域的广泛的PPI接口：两个单体相互作用，形成二聚体通过PPI接口附近的G19。该二聚体然后通过G135附近的PPI界面与另一个单体相互作用以形成三聚体。我们已经发现，扰动G19附近的PPI接口的结果在单体OmpF的形成。设计的二聚体OmpF突变体的热变性表明，寡聚体解离可以从蛋白质解折叠的过程中分离。此外，G57、G59附近的保守位点对于PPI界面是重要的，并且可能为蛋白质-蛋白质相互作用提供必要的支架。
Biogenesis of β-barrel membrane proteins is a complex, multi-step, and as yet incompletely characterized process. The bacterial porin family is perhaps the best studied protein family among the β-barrel membrane proteins that allows diffusion of small solutes across the bacterial outer membrane. In this study, we have identified residues that contribute significantly to the protein-protein interaction (PPI) interface between the chains of Outer Membrane Protein F (OmpF), a trimeric porin, using an empirical energy function in conjunction with an evolutionary analysis. By replacing these residues through site-directed mutagenesis, either with energetically favorable residues or substitutions that do not occur in natural bacterial outer membrane proteins, we succeeded in engineering OmpF mutants with dimeric and monomeric instead of trimeric oligomerization state. Moreover, our results suggest that the oligomerization of OmpF proceeds through a series of interactions involving two distinct regions of the extensive PPI interface: Two monomers interact to form a dimer through the PPI interface near G19. This dimer than interacts with another monomer through the PPI interface near G135 to form a trimer. We have found that perturbing the PPI interface near G19 results in the formation of the monomeric OmpF only. Thermal de-naturation of the designed dimeric OmpF mutant suggests that the oligomer dissociation can be separated from the process of protein unfolding. Furthermore, the conserved site near G57, G59 is important for the PPI interface and might provide the essential scaffold for protein-protein interactions.
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影响因子：
3.4
作者：
Liang, Jie;Naveed, Hammad;Jimenez-Morales, David;Adamian, Larisa;Lin, Meishan
通讯作者：
Lin, Meishan
DOI：
10.1073/pnas.0914229107
发表时间：
2010-05-04
影响因子：
11.1
作者：
Banerjee, Arijit;Mikhailova, Ellina;Bayley, Hagan
通讯作者：
Bayley, Hagan
影响因子：
5.6
作者：
Gessmann D;Mager F;Naveed H;Arnold T;Weirich S;Linke D;Liang J;Nussberger S
通讯作者：
Nussberger S
影响因子：
5.6
作者：
Ben-Shimon, Avraham;Eisenstein, Miriam
通讯作者：
Eisenstein, Miriam
DOI：
10.1002/wnan.50
发表时间：
2009-09
影响因子：
8.6
作者：
Adiga, Shashishekar P.;Jin, Chunmin;Curtiss, Larry A.;Monteiro-Riviere, Nancy A.;Narayan, Roger J.
通讯作者：
Narayan, Roger J.