Erbakan Mustafa, Shen Yue-xiao, Grzelakowski Mariusz, Butler Peter J, Kumar Manish, Curtis Wayne R
Department of Biomedical Engineering, Pennsylvania State University, University Park, Pennsylvania, United States of America.
Department of Chemical Engineering, Pennsylvania State University, University Park, Pennsylvania, United States of America.
PLoS One. 2014 Jan 31;9(1):e86830. doi: 10.1371/journal.pone.0086830. eCollection 2014.
Aquaporins are highly selective water channel proteins integrated into plasma membranes of single cell organisms; plant roots and stromae; eye lenses, renal and red blood cells in vertebrates. To date, only a few microbial aquaporins have been characterized and their physiological importance is not well understood. Here we report on the cloning, expression and characterization of a novel aquaporin, RsAqpZ, from a purple photosynthetic bacterium, Rhodobacter sphaeroides ATCC 17023. The protein was expressed homologously at a high yield (∼20 mg/L culture) under anaerobic photoheterotrophic growth conditions. Stopped-flow light scattering experiments demonstrated its high water permeability (0.17±0.05 cm/s) and low energy of activation for water transport (2.93±0.60 kcal/mol) in reconstituted proteoliposomes at a protein to lipid ratio (w/w) of 0.04. We developed a fluorescence correlation spectroscopy based technique and utilized a fluorescent protein fusion of RsAqpZ, to estimate the single channel water permeability of RsAqpZ as 1.24 (±0.41) x 10(-12) cm(3)/s or 4.17 (±1.38)×10(10) H2O molecules/s, which is among the highest single channel permeability reported for aquaporins. Towards application to water purification technologies, we also demonstrated functional incorporation of RsAqpZ in amphiphilic block copolymer membranes.
水通道蛋白是高度选择性的水通道蛋白，整合到单细胞生物、植物根和基质、晶状体、脊椎动物的肾和红细胞的质膜中。迄今为止，仅对少数微生物水通道蛋白进行了表征，其生理重要性尚未得到充分了解。在此，我们报告了一种来自紫色光合细菌球形红杆菌ATCC 17023的新型水通道蛋白RsAqpZ的克隆、表达和表征。该蛋白在厌氧光异养生长条件下以高产率（约20 mg/L培养物）同源表达。停流光散射实验表明，在蛋白质与脂质比（w/w）为0.04的重组蛋白脂质体中，其具有高水渗透性（0.17±0.05 cm/s）和低水运输活化能（2.93±0.60 kcal/mol）。我们开发了一种基于荧光相关光谱的技术，并利用RsAqpZ的荧光蛋白融合体，估计RsAqpZ的单通道水渗透性为1.24（±0.41）x 10(-12) cm(3)/s或4.17（±1.38）×10(10)个水分子/s，这是报道的水通道蛋白中单通道渗透性最高的之一。为了应用于水净化技术，我们还证明了RsAqpZ在两亲性嵌段共聚物膜中的功能整合。
