Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
J Biol Chem. 2012 Oct 19;287(43):36201-7. doi: 10.1074/jbc.M112.407056. Epub 2012 Sep 6.
The enzymes in the human de novo purine synthesis pathway were found to form a cellular complex, the purinosome, upon culturing cells in purine-depleted medium (An, S., Kumar R., Sheets, E. D., and Benkovic, S. J. (2008) Science 320, 103-106). Purinosome formation and dissociation were found to be modulated by several factors, including the microtubule network and cell signaling involving protein phosphorylation. To determine whether the pathway enzymes are in physical contact, we probed for the protein-protein interactions (PPIs) within the purinosome with a novel application of the Tango PPI reporter system (Barnea, G., Strapps, W., Herrada, G., Berman, Y., Ong, J., Kloss, B., Axel, R., and Lee, K. J. (2008) Proc. Natl. Acad. Sci. U.S.A. 105, 64-69). We found PPIs among all six enzymes within the pathway and evidence for a core involving the first three enzymes. We also captured purinosomes under both purine-rich and purine-depleted conditions. The results provide additional insights into the transient nature and topography of the purinosome.
在嘌呤缺乏的培养基中培养细胞时，人们发现人类从头嘌呤合成途径中的酶会形成一个细胞复合物，即嘌呤体（An, S., Kumar R., Sheets, E. D., and Benkovic, S. J. (2008) Science 320, 103-106）。嘌呤体的形成和解离受到多种因素的调节，包括微管网络和涉及蛋白质磷酸化的细胞信号转导。为了确定途径中的酶是否存在物理接触，我们使用 Tango PPI 报告系统的一种新应用来探测嘌呤体中的蛋白-蛋白相互作用（PPIs）（Barnea, G., Strapps, W., Herrada, G., Berman, Y., Ong, J., Kloss, B., Axel, R., and Lee, K. J. (2008) Proc. Natl. Acad. Sci. U.S.A. 105, 64-69）。我们发现该途径中的所有 6 种酶之间都存在 PPIs，并且有证据表明存在一个涉及前三种酶的核心。我们还在嘌呤丰富和嘌呤缺乏的条件下捕获了嘌呤体。这些结果为嘌呤体的瞬态性质和拓扑结构提供了更多的见解。
