Maurel C, Reizer J, Schroeder J I, Chrispeels M J, Saier M H
Department of Biology, University of California at San Diego, La Jolla 92093-0116.
J Biol Chem. 1994 Apr 22;269(16):11869-72.
The glycerol facilitator of Escherichia coli, GlpF, is a putative nonselective transport channel in the inner membrane of this Gram-negative bacterium. It is a member of the major intrinsic protein (MIP) family of transmembrane channel proteins. Its characterization has been hampered by the lack of a heterologous test system in which its activity can be examined in the absence of other bacterial proteins. Transport of glycerol mediated by this protein was characterized following injection of glpF mRNA into Xenopus laevis oocytes. The properties of GlpF were compared with those of the homologous plant water channel protein, gamma tonoplast intrinsic protein (gamma TIP), as well as the nonhomologous Xenopus K+ channel, Xsha. GlpF selectively transported glycerol but not water or ions, while gamma TIP and Xsha were specific for water and K+, respectively. Voltage clamp experiments showed that GlpF was not voltage-activated for ion transport. Glycerol transport via GlpF proved to be nonsaturable up to 200 mM and exhibited a low temperature of activation (Ea = 4.5 kcal/mol), consistent with the conclusion of Heller et al. (Heller, K. B., Lin, E. C. C., and Wilson, T. H. (1980) J. Bacteriol. 144, 274-278) that GlpF mediates glycerol diffusion via a pore type mechanism. GlpF-mediated transport of glycerol was blocked by mercuric ions (Hg2+) but not N-ethylmaleimide. The inhibitory effect of Hg2+ was partially prevented by inclusion of a high concentration of glycerol and reversed by mercaptoethanol. The results serve to characterize the transport properties of the E. coli glycerol facilitator.
大肠杆菌的甘油转运蛋白GlpF是这种革兰氏阴性细菌内膜中一种推测的非选择性运输通道。它是跨膜通道蛋白主要内在蛋白（MIP）家族的成员。由于缺乏一种能够在没有其他细菌蛋白的情况下检测其活性的异源测试系统，其特性研究受到了阻碍。将glpF mRNA注射到非洲爪蟾卵母细胞后，对该蛋白介导的甘油转运进行了表征。将GlpF的特性与同源植物水通道蛋白γ液泡膜内在蛋白（γTIP）以及非同源的非洲爪蟾钾通道Xsha的特性进行了比较。GlpF选择性转运甘油，而不转运水或离子，而γTIP和Xsha分别对水和钾具有特异性。电压钳实验表明，GlpF对离子转运没有电压激活作用。经GlpF的甘油转运在高达200 mM时被证明是不饱和的，并且表现出较低的活化温度（Ea = 4.5千卡/摩尔），这与Heller等人（Heller, K. B., Lin, E. C. C., and Wilson, T. H. (1980) J. Bacteriol. 144, 274 - 278）得出的GlpF通过孔型机制介导甘油扩散的结论一致。GlpF介导的甘油转运被汞离子（Hg2+）阻断，但不被N - 乙基马来酰亚胺阻断。高浓度甘油的存在可部分阻止Hg2+的抑制作用，巯基乙醇可使其逆转。这些结果有助于表征大肠杆菌甘油转运蛋白的转运特性。
