Institute of Biochemistry and Signal Transduction, University Medical Centre Hamburg-Eppendorf (UKE), Martinistrasse 52, 20246 Hamburg, Germany; Center for Integrated Protein Science Munich (CIPSM), Department Chemistry, Technical University of Munich, Lichtenbergstrasse 4, 85748 Garching, Germany.
Institute of Biochemistry and Signal Transduction, University Medical Centre Hamburg-Eppendorf (UKE), Martinistrasse 52, 20246 Hamburg, Germany; Center for Integrated Protein Science Munich (CIPSM), Department Chemistry, Technical University of Munich, Lichtenbergstrasse 4, 85748 Garching, Germany; Centre for Structural Systems Biology (CSSB), University Medical Centre Hamburg-Eppendorf (UKE), Hamburg, Germany.
Biochim Biophys Acta Proteins Proteom. 2021 Aug;1869(8):140661. doi: 10.1016/j.bbapap.2021.140661. Epub 2021 Apr 16.
Salmonella outer protein D (SopD) is secreted into a host during the first stages of the Salmonella infection and contributes to the systemic virulence of the bacterium. SopD2 is a SopD homolog and possesses GTPase activating protein (GAP) activity towards Rab32. Here, we identified Rab-proteins as putative SopD-targets using a yeast two-hybrid approach. In vitro investigations subsequently revealed Rab8a as an exclusive SopD substrate in contrast to SopD2, which has a broader specificity targeting Rab29, Rab32 and Rab38 in vitro. Additionally, we determined the catalytic efficiencies of SopD and SopD2 towards their physiologically relevant substrates. Moreover, mutagenesis studies provided insights into possible key residues of the Rab-protein and the GAP involved in the conversion of active to inactive GTPase. In conclusion, we demonstrate that Salmonella SopD and SopD2 act as RabGAPs and can inactivate Rab signaling.
沙门氏菌外蛋白 D（SopD）在沙门氏菌感染的早期阶段被分泌到宿主中，并有助于细菌的全身毒力。SopD2 是 SopD 的同源物，对 Rab32 具有 GTP 酶激活蛋白 (GAP) 活性。在这里，我们使用酵母双杂交方法鉴定 Rab 蛋白作为 SopD 的假定靶标。随后的体外研究表明，Rab8a 是 SopD 的唯一底物，而 SopD2 具有更广泛的特异性，可在体外靶向 Rab29、Rab32 和 Rab38。此外，我们确定了 SopD 和 SopD2 对其生理相关底物的催化效率。此外，突变研究深入了解了 Rab 蛋白和 GAP 中可能的关键残基在将活性 GTP 酶转化为非活性 GTP 酶中的作用。总之，我们证明了沙门氏菌 SopD 和 SopD2 作为 RabGAP 起作用，并能使 Rab 信号失活。
