伊利诺斯大学香槟(Urbana-Champaign)分校和亚利桑那大学的研究人员将计算机模拟手段和实验室技术联合起来,发现aquaporins(水通道蛋白)发挥门控机制(gating mechanism)的一种关键亚基——loop D,不仅控制水分子的出入而且与离子传导有关,发挥哪种功能主要取决于刺激细胞的信号途径。研究结果发表于《Structure》9月刊。
Aquaporin是一系列能够控制水分子在细胞和细胞外环境间传递的膜蛋白组成的家族。伊利诺斯大学香槟(Urbana-Champaign)分校的研究人员和亚利桑那大学的研究人员发现此家族的成员之一,aquaporin-1也具有离子通道(ion channels)的功能。
伊利诺斯大学生化教授、Beckman 尖端科技研究所研究员Emad Tajkhorshid说:“弄清门控通道的分子机制,有助于蛋白工程学研究。”
Tajkhorshid等利用已知的aquaporin-1晶体结构和已掌握的分子动力学模拟(molecular dynamics simulations)技术,发现aquaporin-1的中心孔道(central pore,生物通编者译)能够传递,其门控功能受到细胞内环磷酸鸟苷(cGMP或cyclic GMP或3'-5'-cyclic guanosine monophosphate)信号控制。cGMP诱导aquaporin的中心孔道的 loop D构象改变。
“loop D环非常柔韧,每排中有四个带正电荷的精氨酸残基,并且有的精氨酸残基会延伸到中心孔道,”Tajkhorshid说,“我们发现cGMP与loop D结合,促进了精氨酸残基向细胞外的运动,刺激了门道的打开。”
研究人员高度强调了将模拟和实验结合使用的优势。利用模拟技术,研究人员设计了一种突变模型,模型中两个精氨酸被两个丙胺酸(Alanines)取代。在Arizona实验室进行的实验证明,这种突变通道的导电功能几乎消失了,但是输水功能不受影响。
“弄清这种机制有助于我们更好地控制中心孔道的开启和关闭” Tajkhorshid说,“通过修饰孔道中的残基或者改变控制门控孔道的loop D的长度,我们可以关闭传导离子的功能,或者设计出新的开启更为方便、导电率更高的水通道蛋白。”
研究受到美国国立卫生研究院资助。合作者包括:伊利诺斯大学Jin Yu(于金,音译)、Klaus Schulten和亚利桑那大学Andrea J. Yool。
英文原文:
One protein, two channels: Scientists explain mechanism in aquaporins
Using computer simulations and experimental results, researchers at the University of Illinois at Urbana-Champaign and the University of Arizona have identified a key component of the gating mechanism in aquaporins that controls both the passage of water and the conduction of ions.
Aquaporins are a class of proteins that form membrane channels in cell walls and allow for water movement between a cell and its surroundings. A number of aquaporins, including aquaporin-1, have been found to function as ion channels, as well.
“Understanding the molecular mechanism behind gating in membrane channels could lead to more effective protein engineering,” said Emad Tajkhorshid, a professor of biochemistry at Illinois and a researcher at the Beckman Institute for Advanced Science and Technology.
In work funded by the National Institutes of Health, Tajkhorshid and co-workers show that the same protein can be used as a water channel or an ion channel depending on the signaling pathway activated in the cell. The scientists report their findings in the September issue of the journal Structure.
Taking advantage of the known crystal structure of aquaporin-1 and the power of molecular dynamics simulations, the researchers explored the central pore as a candidate pathway for conducting ions. Gating of the central pore is controlled by cyclic guanosine monophosphate, a signaling nucleotide inside the cell, which induces a conformational change in one of the aquaporin loops (loop D).
“This loop is very flexible, has four positively charged arginine residues in a row, and extends into the central pore,” Tajkhorshid said. “We believe the cGMP interacts with loop D, facilitating its outward motion, which triggers the opening of the gate.”
The work highlights a close interaction between simulation and experiment. Based on their simulation results, the researchers designed a mutant in which two arginines in loop D were replaced by two alanines. In laboratory experiments performed at Arizona, the substitution caused an almost complete removal of ion conduction, but had no appreciable effect on water passage.
“Knowing the mechanism gives us a new handle to control the opening or closing of the central pore,” Tajkhorshid said. “By modifying the pore-lining residue, or altering the length of loop D that gates the pore, we can shut down the ion conductivity completely, or engineer new aquaporins that can be opened more easily or have a higher ion conduction rate once open.”
With Tajkhorshid, co-authors of the paper are Illinois graduate student and lead author Jin Yu, University of Arizona experimentalist Andrea J. Yool, and Illinois physicist Klaus Schulten.