Pseudomonas aeruginosa porin OprF -: Properties of the channel
Pseudomonas aeruginosa porin OprF -: Properties of the channel
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DOI：
10.1074/jbc.m600650200
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发表时间：
2006-06-16
影响因子：
4.8
通讯作者：
Bezrukov, Sergey M.
中科院分区：
文献类型：
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作者：
Nestorovich, Ekaterina M.;Sugawara, Etsuko;Bezrukov, Sergey M.
利用离子通道重构技术，我们研究了铜绿假单胞菌OprF亚组分在两种不同构象中的通道形成活性：少数单域构象和多数双域构象(Sugawa，E.，Nestorovich，E.M.，Bezrukov，S.M.和Nikaido，H.(2006)J.Biol)。化学。281,16220 16229)。由于单区构象的部分被耗尽，我们无法检测到任何具有明确电导水平的通道的形成。对于普通的OprF，我们只看到了罕见的通道形成。然而，对于OprF的单一结构域富集组分，我们观察到具有高度重复性的电导的有规律的插入。单一的OprF通道表现出丰富的动力学行为，表现出在聚合物排斥实验中测量到的不同离子电导和半径的几个亚构象之间的自发跃迁。虽然我们发现大多数导电构象的有效半径超过了大肠杆菌的一般外膜孔道OmpF的有效半径，但我们也发现单一的OprF通道主要存在于弱导电的亚构象中，并且只在很短的时间内切换到全开放状态。因此，早期报道的OprF的低渗透性可能是由于两个因素：主要是由于OprF群体中单域构象的稀少，其次是由于单域构象中弱导电亚构象的优势。
Using ion channel reconstitution in planar lipid bilayers, we examined the channel-forming activity of subfractions of Pseudomonas aeruginosa OprF, which was shown to exist in two different conformations: a minority single domain conformer and a majority two-domain conformer ( Sugawara, E., Nestorovich, E. M., Bezrukov, S. M., and Nikaido, H. ( 2006) J. Biol. Chem. 281, 16220 16229). With the fraction depleted for the single domain conformer, we were unable to detect formation of any channels with well defined conductance levels. With the unfractionated OprF, we saw only rare channel formation. However, with the single domain-enriched fraction of OprF, we observed regular insertion of channels with highly reproducible conductances. Single OprF channels demonstrate rich kinetic behavior exhibiting spontaneous transitions between several subconformations that differ in ionic conductance and radius measured in polymer exclusion experiments. Although we showed that the effective radius of the most conductive conformation exceeds that of the general outer membrane porin of Escherichia coli, OmpF, we also found that a single OprF channel mainly exists in weakly conductive subconformations and switches to the fully open state for a short time only. Therefore, the low permeability of OprF reported earlier may be due to two factors: mainly to the paucity of the single domain conformer in the OprF population and secondly to the predominance of weakly conductive subconformations within the single domain conformer.