Three two-component transporters with channel-like properties have monovalent cation/proton antiport activity
Three two-component transporters with channel-like properties have monovalent cation/proton antiport activity
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DOI：
10.1073/pnas.0703709104
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发表时间：
2007-08-14
影响因子：
11.1
通讯作者：
Krulwich, Terry A.
中科院分区：
文献类型：
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作者：
Fujisawa, Makoto;Ito, Masahiro;Krulwich, Terry A.
阳离子/质子反向转运蛋白-2（CPA 2）家族的四种双组分细菌转运系统的性质导致了这样的建议，即该CPA 2亚组可以使用通道而不是反向转运机制[参见Booth IR，Edwards MD，Gunasekera B，Li C，米勒S（2005）in Bacterial Ion Channels，eds Kubalski A，Martinac B（Am Soc Microbiol，华盛顿，DC）第21-40页]。转运子亚组包括深入研究的谷胱甘肽门控的K+外排系统从大肠杆菌，KefGB和KefFC。KefG和KefF是辅助蛋白。它们是在操纵子中与各自的转运蛋白Kef B和KefC一起编码的外周膜蛋白，并且是最佳外排活性所需的。该亚组的其他双组分CPA 2转运蛋白是AmhMT，一种来自嗜碱假坚强芽孢杆菌OF 4的NH 4+（K+）流出系统;和YhaTU，一种来自枯草芽孢杆菌的K+流出系统。在此，证明了来自反向转运蛋白缺陷型E. coli KNabc. K+的表观Km在低mM范围内。外周蛋白是YhaU和KefC依赖性反向转运所必需的，而AmhT和AmhMT都表现出反向转运。KefFC具有最宽的底物范围，使用Rb+近似于K+> Li+> Na+。Glucoside显著抑制KefFC介导的囊泡中的K+/H+反向转运。抑制作用被NADH增强，推测其与KefC的KTN/RCK结构域结合。反向转运机制解释了体内KefFC介导的亲电体抗性中涉及的H+摄取。由于AmhMT在嗜碱菌中的生理底物是NH 4+，因此该结果还暗示AmhMT催化NH 4 +/H+反向转运，这将阻止NH 4+流出过程中细胞质H+的净损失。
Properties of four two-component bacterial transport systems of the cation/proton antiporter-2 (CPA2) family led to suggestions that this CPA2 subset may use a channel rather than an antiport mechanism [see Booth IR, Edwards MD, Gunasekera B, Li C, Miller S (2005) in Bacterial Ion Channels, eds Kubalski A, Martinac B (Am Soc Microbiol, Washington, DC) pp 21-40]. The transporter subset includes the intensively studied glutathione-gated K+ efflux systems from Escherichia coli, KefGB, and KefFC. KefG and KefF are ancillary proteins. They are peripheral membrane proteins that are encoded in operons with the respective transporter proteins, Kef B and KefC, and are required for optimal efflux activity. The other two-component CPA2 transporters of the subset are AmhMT, an NH4+ (K+) efflux system from alkaliphilic Bacillus pseudofirmus OF4; and YhaTU, a K+ efflux system from Bacillus subtilis. Here a K+/H+ antiport capacity was demonstrated for YhaTU, AmhMT, and KefFC in membrane vesicles from antiporter-deficient E. coli KNabc. The apparent Km for K+ was in the low mM range. The peripheral protein was required for YhaU- and KefC-dependent antiport, whereas both AmhT and AmhMT exhibited antiport. KefFC had the broadest range of substrates, using Rb+approximate to K+> Li+> Na+. Glutathione significantly inhibited KefFC-mediated K+/H+ antiport in vesicles. The inhibition was enhanced by NADH, which presumably binds to the KTN/RCK domain of KefC. The antiport mechanism accounts for the H+ uptake involved in KefFC-mediated electrophile resistance in vivo. Because the physiological substrate of AmhMT in the alkaliphile is NH4+, the results also imply that AmhMT catalyzes NH4+/H+ antiport, which would prevent net cytoplasmic H+ loss during NH4+ efflux.