Eukaryotic cell determination of ExoS ADP-ribosyltransferase substrate specificity
Eukaryotic cell determination of ExoS ADP-ribosyltransferase substrate specificity
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DOI：
10.1006/bbrc.2002.6402
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发表时间：
2002-02-15
影响因子：
3.1
通讯作者：
Olson, JC
中科院分区：
文献类型：
--
作者：
Fraylick, JE;Rucks, EA;Olson, JC
Exoenzyme S（ExoS）ADP-核糖基化体外多种低分子量G-（LMMG-）蛋白。ExoS的体内底物特异性的鉴定受到其细菌接触递送到真核细胞中以及鉴定细胞内ADP-核糖基化蛋白质的困难的阻碍。通过二维电泳比较ExoS在体外对底物的修饰与细菌易位到HT-29上皮细胞中后的修饰，将Ras、Ral和Rab蛋白和Rac 1鉴定为ExoS ADPRT活性的体内底物。细胞分级分离研究确定了膜缔合和底物修饰效率之间的关系。此外，Rac和Cdc 42响应于ExoS而重新定位于膜。底物修饰与暴露于ExoS的时间的比较鉴定了底物修饰的进展，首先修饰Ras、RalA和Rab 5，然后是RabS和11，然后是Rab 7和Rac 1。数据支持LMMG蛋白的内在性质及其亚细胞定位是细菌易位ExoS底物选择性的决定因素。(C)2002 Elsevier Science（美国）。
Exoenzyme S (ExoS) ADP-ribosylates multiple low-molecular-mass G- (LMMG-) proteins in vitro. Identification of the in vivo substrate specificity of ExoS has been hindered by its bacterial contact delivery into eukaryotic cells and difficulties in identifying ADP-ribosylated proteins within cells. Two-dimensional electrophoresis comparisons of substrate modifications by ExoS in vitro to that following bacterial translocation into HT-29 epithelial cells identified Ras, Ral, and Rab proteins and Rac1 as in vivo substrates of ExoS ADPRT activity. Cellular fractionation studies identified a relationship between membrane association and efficiency of substrate modification. Moreover, Rac and Cdc42 relocalized to the membrane in response to ExoS. Comparisons of substrate modification to time of exposure to ExoS identified a progression of substrate modification, with Ras, RalA, and Rab5 modified first, followed by RabS and 11, then Rab7 and Rac1. The data support that intrinsic properties of LMMG-proteins and their subcellular localization are determinants of bacterially translocated ExoS substrate selectivity. (C) 2002 Elsevier Science (USA).