Fotiadis D, Jenö P, Mini T, Wirtz S, Müller S A, Fraysse L, Kjellbom P, Engel A
M. E. Müller-Institute for Microscopy, Biozentrum of the University of Basel, CH-4056 Basel, Switzerland.
J Biol Chem. 2001 Jan 19;276(3):1707-14. doi: 10.1074/jbc.M009383200. Epub 2000 Oct 24.
Two members of the aquaporin family, PM28A and a new one, PM28C, were isolated and shown to be the major constituents of spinach leaf plasma membranes. These two isoforms were identified and characterized by matrix-assisted laser desorption ionization-mass spectrometry. Edman degradation yielded the amino acid sequence of two domains belonging to the new isoform. PM28B, a previously described isoform, was not found in our preparations. Scanning transmission electron microscopy mass analysis revealed both PM28 isoforms to be tetrameric. Two types of particles, a larger and a smaller one, were found by transmission electron microscopy of negatively stained solubilized proteins and by atomic force microscopy of PM28 two-dimensional crystals. The ratio of larger to smaller particles observed by transmission electron microscopy and single particle analysis correlated with the ratio of PM28A to PM28C determined by matrix-assisted laser desorption ionization-mass spectrometry. The absence of PM28B and the ratio of PM28A to PM28C indicate that these plasma membrane intrinsic proteins are differentially expressed in spinach leaves. These findings suggest that differential expression of the various aquaporin isoforms may regulate the water flux across the plasma membrane, in addition to the known mechanism of regulation by phosphorylation.
水通道蛋白家族的两个成员，PM28A和一个新成员PM28C，被分离出来并被证明是菠菜叶质膜的主要成分。这两种异构体通过基质辅助激光解吸电离质谱法进行鉴定和表征。埃德曼降解法得到了属于新异构体的两个结构域的氨基酸序列。在我们的制备物中未发现先前描述的异构体PM28B。扫描透射电子显微镜质量分析表明两种PM28异构体均为四聚体。通过对负染溶解蛋白的透射电子显微镜观察以及对PM28二维晶体的原子力显微镜观察，发现了两种类型的颗粒，一种较大，一种较小。通过透射电子显微镜和单颗粒分析观察到的较大颗粒与较小颗粒的比例，与通过基质辅助激光解吸电离质谱法测定的PM28A与PM28C的比例相关。PM28B的缺失以及PM28A与PM28C的比例表明，这些质膜内在蛋白在菠菜叶中差异表达。这些发现表明，除了已知的磷酸化调节机制外，各种水通道蛋白异构体的差异表达可能调节跨质膜的水通量。
