Schröder S, Lohse M J
Institute of Pharmacology, University of Würzburg, Germany.
Naunyn Schmiedebergs Arch Pharmacol. 2000 Nov;362(4-5):435-9. doi: 10.1007/s002100000298.
Phosducin-like protein is a protein with wide-spread expression that has been shown to be capable of inhibiting G-protein function in vitro. However, it is not clear whether it is expressed in sufficient amounts to actually exert such functions in vivo. Here we quantify the expression of the short and the long splice variants of phosducin-like protein, PhlPs and PhlP1. Western blots of various rat tissues showed that PhlP1 was by far the dominant splice variant; its levels were 1.5-2 pmol/mg cytosolic protein in brain, liver and kidney, and about 0.5 pmol/mg cytosolic protein in lung, heart and skeletal muscle. These values correspond to concentrations of 150-200 nM and 50 nM, respectively. The levels of PhlPs were about 20-fold lower. Recombinant phosducin, PhlP1 and PhlPs inhibited the interaction between G-protein alpha- und betagamma-subunits with IC50-values of 6 nM, 6 nM and 90 nM, respectively, as determined by Gbetagamma-dependent ADP-ribosylation of Galphai1 by pertussis-toxin. Thus, tissue concentrations of PhlP1 are clearly sufficient to affect G-protein function in vivo, while the expression levels and the Gbetagamma-affinity of PhlPs are most likely too low to have significant inhibitory effects on Gbetagamma (G-protein betagamma-subunits).
类磷光蛋白是一种广泛表达的蛋白质，已证明其在体外能够抑制G蛋白功能。然而，尚不清楚其在体内的表达量是否足以实际发挥此类功能。在此，我们对类磷光蛋白的短剪接变体和长剪接变体PhlPs和PhlP1的表达进行了定量。对各种大鼠组织进行的蛋白质免疫印迹分析表明，PhlP1是迄今为止主要的剪接变体；其在脑、肝和肾中的水平为1.5 - 2 pmol/mg胞质蛋白，在肺、心脏和骨骼肌中的水平约为0.5 pmol/mg胞质蛋白。这些值分别对应于150 - 200 nM和50 nM的浓度。PhlPs的水平低约20倍。通过百日咳毒素对Gαi1进行Gβγ依赖性ADP核糖基化测定，重组磷光蛋白、PhlP1和PhlPs抑制G蛋白α亚基和βγ亚基之间相互作用的IC50值分别为6 nM、6 nM和90 nM。因此，PhlP1的组织浓度显然足以在体内影响G蛋白功能，而PhlPs的表达水平和对Gβγ的亲和力很可能过低，无法对Gβγ（G蛋白βγ亚基）产生显著抑制作用。
