Maierhofer Tobias, Lind Christof, Hüttl Stefanie, Scherzer Sönke, Papenfuß Melanie, Simon Judy, Al-Rasheid Khaled A S, Ache Peter, Rennenberg Heinz, Hedrich Rainer, Müller Thomas D, Geiger Dietmar
University of Würzburg, Institute for Molecular Plant Physiology and Biophysics, D-97082 Würzburg, Germany.
Institute of Forest Sciences, Chair of Tree Physiology, University of Freiburg, 79110 Freiburg, Germany.
Plant Cell. 2014 Jun;26(6):2554-2567. doi: 10.1105/tpc.114.125849. Epub 2014 Jun 17.
In contrast to animal cells, plants use nitrate as a major source of nitrogen. Following the uptake of nitrate, this major macronutrient is fed into the vasculature for long-distance transport. The Arabidopsis thaliana shoot expresses the anion channel SLOW ANION CHANNEL1 (SLAC1) and its homolog SLAC1 HOMOLOGOUS3 (SLAH3), which prefer nitrate as substrate but cannot exclude chloride ions. By contrast, we identified SLAH2 as a nitrate-specific channel that is impermeable for chloride. To understand the molecular basis for nitrate selection in the SLAH2 channel, SLAC1 and SLAH2 were modeled to the structure of HiTehA, a distantly related bacterial member. Structure-guided site-directed mutations converted SLAC1 into a SLAH2-like nitrate-specific anion channel and vice versa. Our findings indicate that two pore-occluding phenylalanines constrict the pore. The selectivity filter of SLAC/SLAH anion channels is determined by the polarity of pore-lining residues located on alpha helix 3. Changing the polar character of a single amino acid side chain (Ser-228) to a nonpolar residue turned the nitrate-selective SLAH2 into a chloride/nitrate-permeable anion channel. Thus, the molecular basis of the anion specificity of SLAC/SLAH anion channels seems to be determined by the presence and constellation of polar side chains that act in concert with the two pore-occluding phenylalanines.
与动物细胞不同，植物将硝酸盐作为主要的氮源。硝酸盐被吸收后，这种主要的大量营养素会进入脉管系统进行长距离运输。拟南芥地上部分表达阴离子通道慢阴离子通道1（SLAC1）及其同源物慢阴离子通道1同源物3（SLAH3），它们优先选择硝酸盐作为底物，但不能排除氯离子。相比之下，我们鉴定出SLAH2是一种对氯离子不通透的硝酸盐特异性通道。为了理解SLAH2通道中硝酸盐选择的分子基础，我们根据一种远缘相关细菌成员HiTehA的结构对SLAC1和SLAH2进行了建模。基于结构的定点突变将SLAC1转化为类似SLAH2的硝酸盐特异性阴离子通道，反之亦然。我们的研究结果表明，两个堵塞孔道的苯丙氨酸会使孔道变窄。SLAC/SLAH阴离子通道的选择性过滤器由位于α螺旋3上的孔道内衬残基的极性决定。将单个氨基酸侧链（Ser-228）的极性特征改变为非极性残基，会使硝酸盐选择性的SLAH2变成一种可通透氯离子/硝酸盐的阴离子通道。因此，SLAC/SLAH阴离子通道阴离子特异性的分子基础似乎由极性侧链的存在和排列决定，这些极性侧链与两个堵塞孔道的苯丙氨酸协同作用。
