Structural basis for dynamic mechanism of nitrate/nitrite antiport by NarK.
Structural basis for dynamic mechanism of nitrate/nitrite antiport by NarK.
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DOI：
10.1038/ncomms8097
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发表时间：
2015-05-11
影响因子：
16.6
通讯作者：
Nureki, Osamu
中科院分区：
文献类型：
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作者：
Fukuda, Masahiro;Takeda, Hironori;Kato, Hideaki E.;Doki, Shintaro;Ito, Koichi;Maturana, Andres D.;Ishitani, Ryuichiro;Nureki, Osamu
NarK属于硝酸盐/亚硝酸盐转运蛋白（NNP）家族，在主要易化剂超家族（MFS）中，并在多种生物体（包括古细菌，细菌，真菌和植物）的跨膜硝酸盐吸收中发挥核心作用。虽然以前的研究提供了深入了解的整体结构和底物识别的NarK，其分子机制，包括硝酸盐运输的驱动力，仍然难以捉摸。在这里，我们证明了NarK是一个硝酸盐/亚硝酸盐反向转运蛋白，使用体外重建系统。此外，我们提出了高分辨率的晶体结构的NarK从大肠杆菌在硝酸盐结合闭塞，硝酸盐结合向内开放和apo向内开放状态。综合的结构，功能和计算分析揭示了硝酸盐/亚硝酸盐反向转运机制的NarK，其中底物识别耦合到运输循环的跨膜螺旋和关键的酪氨酸和精氨酸残基在底物结合位点的伴随运动。 硝酸盐/亚硝酸盐转运蛋白（NNP）在古菌、细菌、真菌和植物的硝酸盐吸收过程中起着重要作用。在这里，Fukuda等人使用基于脂质体的转运测定，X射线晶体学和分子动力学模拟来揭示细菌NNP，NarK的动态硝酸盐/亚硝酸盐反向转运机制。
NarK belongs to the nitrate/nitrite porter (NNP) family in the major facilitator superfamily (MFS) and plays a central role in nitrate uptake across the membrane in diverse organisms, including archaea, bacteria, fungi and plants. Although previous studies provided insight into the overall structure and the substrate recognition of NarK, its molecular mechanism, including the driving force for nitrate transport, remained elusive. Here we demonstrate that NarK is a nitrate/nitrite antiporter, using an in vitro reconstituted system. Furthermore, we present the high-resolution crystal structures of NarK from Escherichia coli in the nitrate-bound occluded, nitrate-bound inward-open and apo inward-open states. The integrated structural, functional and computational analyses reveal the nitrate/nitrite antiport mechanism of NarK, in which substrate recognition is coupled to the transport cycle by the concomitant movement of the transmembrane helices and the key tyrosine and arginine residues in the substrate-binding site. Nitrate/nitrite porters (NNP) play a central role in nitrate uptake in archaea, bacteria, fungi and plants. Here, Fukuda et al. use a liposome-based transport assay, X-ray crystallography and molecular dynamics simulation to reveal the dynamic nitrate/nitrite antiport mechanism of a bacterial NNP, NarK.
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影响因子：
3.3
作者：
Klauda, Jeffery B.;Venable, Richard M.;Freites, J. Alfredo;O'Connor, Joseph W.;Tobias, Douglas J.;Mondragon-Ramirez, Carlos;Vorobyov, Igor;MacKerell, Alexander D., Jr.;Pastor, Richard W.
通讯作者：
Pastor, Richard W.
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
DOI：
10.1107/s0907444992007698
发表时间：
1993-01-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
COWTAN, KD;MAIN, P
通讯作者：
MAIN, P
影响因子：
3.9
作者：
Maria Martinez-Espinosa, Rosa;Cole, Jeffrey A.;Watmough, Nicholas J.
通讯作者：
Watmough, Nicholas J.
大肠杆菌K-12的构造框架，单基因敲除突变体：Keio Collection。
影响因子：
9.9
作者：
通讯作者：
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