Structure-based analysis of biological activity of Helicobacter pylori CagA

幽门螺杆菌CagA蛋白被注射到胃上皮细胞中，在胃上皮细胞中，它通过其含有Glu-Pro-Ile-Tyr-Ala（EPIYA）基序的C-末端区域与细胞蛋白如SHP-2和Par 1相互作用。因此，CagA干扰参与细胞生长和细胞极性调节的细胞内机制。为了阐明CagA的病理生理活性的分子基础，我们试图确定CagA的三维结构。我们进行了有限的CagA蛋白水解实验，并确定了一个特定的切割位点，这可能是一个连接区，连接可能的N-末端和C-末端结构域的CagA。有趣的是，CagA的C-末端蛋白水解片段与剩余的N-末端蛋白水解片段结合。此外，我们发现N-末端片段增强CagA的生物活性，诱导细胞形态学变化依赖于CagA的C-末端区域。我们的研究结果表明，分子内的N-末端和C-末端结构域之间的相互作用起着重要的作用，在生物活性的CagA蛋白的结构完整性。

Helicobacter pylori CagA protein is injected into gastric epithelial cells, where it interacts with cellular proteins such as SHP-2 and Par1 through its C-terminal region containing Glu-Pro-Ile-Tyr-Ala (EPIYA) motifs. Consequently, CagA perturbs intracellular machineries involved in the regulation of cell growth and cell polarity. To elucidate molecular basis for the pathophysiological activity of CagA, we sought to determine three-dimensional structure of CagA. We performed a limited proteolysis experiment of CagA and identified a specific cleavage site, which could represent a linker region that connects the possible N-terminal and C-terminal structural domains of CagA. Intriguingly, the C-terminal proteolytic fragment of CagA was bound to the remaining N-terminal proteolytic fragment. Furthermore, we found that N-terminal fragment enhances CagA biological activity that induces cell morphological changes in depending on the C-terminal region of CagA. Our finding indicates that an intramolecular interaction between the N-terminal and C-terminal domains plays an important role in the structural integrity of the biologically active CagA protein.