Laboratoire de Phosphorylation des Protéines et Pathogénie Bactérienne, Institut de Biologie et Chimie des Protéines, UMR 5086-CNRS/Université de Lyon, IFR128 BioSciences Gerland Lyon Sud, 7 Passage du Vercors, F-69367 Lyon cedex 07, France.
J Mol Biol. 2010 Dec 17;404(5):847-58. doi: 10.1016/j.jmb.2010.10.012. Epub 2010 Oct 19.
PASTA (penicillin-binding protein and serine/threonine kinase associated) modules are found in penicillin-binding proteins and bacterial serine/threonine kinases mainly from Gram-positive Firmicutes and Actinobacteria. They may act as extracellular sensors by binding peptidoglycan fragments. We report here the first crystal structure of a multiple-PASTA domain from Ser/Thr kinase, that of the protein serine/threonine kinase 1 (Stk1) from the Firmicute Staphylococcus aureus. The extended conformation of the three PASTA subunits differs strongly from the compact conformation observed in the two-PASTA domain of penicillin-binding protein PBP2x, whereas linear conformations were also reported for two-subunit fragments of the four-PASTA domain of the Actinobacteria Mycobacterium tuberculosis studied by liquid NMR. Thus, a stretched organization appears to be the signature of modular PASTA domains in Ser/Thr kinases. Signal transduction to the kinase domain is supposed to occur via dimerization and ligand binding. A conserved X-shaped crystallographic dimer stabilized by intermolecular interactions between the second PASTA subunits of each monomer is observed in the two crystal forms of Stk1 that we managed to crystallize. Extracellular PASTA domains are composed of at least two subunits, and this molecular assembly is a plausible candidate for the biological dimer. We have also performed docking experiments, which predict that the hinge regions of the PASTA domain can accommodate peptidoglycan. Finally, a three-dimensional homology molecular model of full-length Stk1 was generated, suggesting an interaction between the kinase domain and the cytoplasmic face of the plasma membrane via a eukaryotic-like juxtamembrane domain. A comprehensive activation mechanism for bacterial Ser/Thr kinases is proposed with the support of these structural data.
PASTA（青霉素结合蛋白和丝氨酸/苏氨酸激酶相关）模块存在于青霉素结合蛋白和细菌丝氨酸/苏氨酸激酶中，主要来自革兰阳性Firmicutes 和 Actinobacteria。它们可以通过结合肽聚糖片段作为细胞外传感器。我们在这里报告了来自Firmicute 金黄色葡萄球菌的 Ser/Thr 激酶蛋白丝氨酸/苏氨酸激酶 1（Stk1）的首个多 PASTA 结构域的晶体结构。三个 PASTA 亚基的伸展构象与在青霉素结合蛋白 PBP2x 的两个 PASTA 结构域中观察到的紧凑构象有很大不同，而在通过液体 NMR 研究的 Actinobacteria 分枝杆菌的四 PASTA 结构域的两个亚基片段中也报道了线性构象。因此，伸展的组织似乎是 Ser/Thr 激酶中模块化 PASTA 结构域的特征。信号转导到激酶结构域被认为是通过二聚化和配体结合发生的。在我们设法结晶的两种 Stk1 晶体形式中观察到由单体的每个分子的第二个 PASTA 亚基之间的分子间相互作用稳定的保守 X 形晶体学二聚体。细胞外 PASTA 结构域由至少两个亚基组成，这种分子组装是生物二聚体的合理候选者。我们还进行了对接实验，预测 PASTA 结构域的铰链区域可以容纳肽聚糖。最后，生成了全长 Stk1 的三维同源分子模型，表明激酶结构域通过类似于真核的跨膜结构域与质膜的细胞质面相互作用。这些结构数据支持提出了细菌 Ser/Thr 激酶的全面激活机制。
