Substrate Specificity of the FurE Transporter Is Determined by Cytoplasmic Terminal Domain Interactions
Substrate Specificity of the FurE Transporter Is Determined by Cytoplasmic Terminal Domain Interactions
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FurE 转运蛋白的底物特异性由细胞质末端结构域相互作用决定
DOI：
10.1534/genetics.117.300327
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发表时间：
2017
期刊：
影响因子：
3.3
通讯作者：
G. Diallinas
中科院分区：
文献类型：
--
作者：
G. Papadaki;S. Amillis;G. Diallinas
Fure是核酸碱基转运体1转运体家族的成员，对尿囊素、尿酸(UA)、尿嘧啶及相关类似物具有特异性。在这里，我们证明了C-或N-末端截短的Fure转运蛋白(Fure-ΔC或Fure-ΔΝ)表现出更高的蛋白质稳定性，但也不能进行UA转运。为了更好地了解细胞质末端区域的作用，我们描述了恢复Fure-ΔC介导的UA运输的基因抑制物。抑制子定位于底物结合位点的外围[跨膜片段(TMS)3中的Thr133和TM 8中的Val343]、外向门(TM 7中的Ser296、TM 9中的Ile371、以及TM 10中的Tyr392和Leu394)，或在柔性环中(LN中的Asp26、L5中的Gly222和L7中的Asn308)。选择的抑制子也被证明恢复了Fure-ΔΝ的野生型特异性，表明C-末端和/或N-末端结构域都参与了对底物选择至关重要的分子内动力学。双分子荧光互补分析支持直接底物敏感的C-末端和/或N-末端结构域的相互作用。据我们所知，这是第一例不仅真核转运蛋白的功能，而且特异性，由其末端细胞质区域调节。
FurE, a member of the Nucleobase Cation Symporter 1 transporter family in Aspergillus nidulans, is specific for allantoin, uric acid (UA), uracil, and related analogs. Herein, we show that C- or N-terminally-truncated FurE transporters (FurE-ΔC or FurE-ΔΝ) present increased protein stability, but also an inability for UA transport. To better understand the role of cytoplasmic terminal regions, we characterized genetic suppressors that restore FurE-ΔC-mediated UA transport. Suppressors map in the periphery of the substrate-binding site [Thr133 in transmembrane segment (TMS)3 and Val343 in TMS8], an outward-facing gate (Ser296 in TMS7, Ile371 in TMS9, and Tyr392 and Leu394 in TMS10), or in flexible loops (Asp26 in LN, Gly222 in L5, and Asn308 in L7). Selected suppressors were also shown to restore the wild-type specificity of FurE-ΔΝ, suggesting that both C- and/or N-terminal domains are involved in intramolecular dynamics critical for substrate selection. A direct, substrate-sensitive interaction of C- and/or N-terminal domains was supported by bimolecular fluorescence complementation assays. To our knowledge, this is the first case where not only the function, but also the specificity, of a eukaryotic transporter is regulated by its terminal cytoplasmic regions.
影响因子：
3.9
作者：
Alguel Y;Cameron AD;Diallinas G;Byrne B
通讯作者：
Byrne B
影响因子：
--
作者：
Colas C;Ung PM;Schlessinger A
通讯作者：
Schlessinger A