Pucciarelli M Graciela, García-del Portillo Francisco
Departamento de Biotecnología Microbiana, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CSIC), Campus de Cantoblanco, 28049 Madrid, Spain.
Mol Microbiol. 2003 Apr;48(2):573-85. doi: 10.1046/j.1365-2958.2003.03469.x.
The invasion-associated type III secretion system of Salmonella enterica assembles as a supra-molecular structure, termed needle complex, which spans the bacterial envelope. Here, we present evidence for protein-peptidoglycan interactions that modulate the assembly of this organelle. The presence of major membrane components of the needle complex (PrgH, PrgK and InvG) and InvH, required for efficient assembly of the organelle, was examined in peptidoglycan purified by extensive boiling of bacteria in 4% SDS. InvH, PrgH and PrgK, but not InvG, were detected in this purified material. InvH was present in the peptidoglycan in higher relative amounts than PrgH or PrgK, and was the only protein efficiently bound to peptidoglycan in cross-linking experiments. Analysis in mutants defective for needle complex proteins showed that the needle proteins PrgI and PrgJ and, to a lesser extent, InvH, sustain the association of PrgH and PrgK with peptidoglycan. In contrast, the association of InvH with peptidoglycan did not necessitate other needle complex proteins. Functional analysis showed that the association of InvH, PrgH and PrgK with peptidoglycan is abolished in live bacteria carrying structural modifications in the peptidoglycan. The loss of these interactions caused a marked reduction in the number of needle complexes and, concomitantly, in protein secretion and bacterial invasion of cultured eukaryotic cells. Altogether, these data provide the first evidence for an association between proteins of the Salmonella needle complex and the peptidoglycan. In addition, we demonstrate that these protein-peptidoglycan interactions are critical for an efficient and correct assembly of this specialized organelle.
肠炎沙门氏菌的侵袭相关III型分泌系统组装成一种超分子结构，称为针状复合体，它跨越细菌包膜。在此，我们提供了蛋白质与肽聚糖相互作用调节该细胞器组装的证据。在通过将细菌在4%十二烷基硫酸钠中长时间煮沸纯化得到的肽聚糖中，检测了针状复合体的主要膜成分（PrgH、PrgK和InvG）以及该细胞器高效组装所需的InvH的存在情况。在这种纯化材料中检测到了InvH、PrgH和PrgK，但未检测到InvG。InvH在肽聚糖中的相对含量高于PrgH或PrgK，并且是交联实验中唯一能有效结合到肽聚糖上的蛋白质。对针状复合体蛋白缺陷型突变体的分析表明，针状蛋白PrgI和PrgJ，以及在较小程度上的InvH，维持了PrgH和PrgK与肽聚糖的结合。相比之下，InvH与肽聚糖的结合并不需要其他针状复合体蛋白。功能分析表明，在肽聚糖中携带结构修饰的活细菌中，InvH、PrgH和PrgK与肽聚糖的结合被消除。这些相互作用的丧失导致针状复合体数量显著减少，同时蛋白质分泌和培养的真核细胞的细菌侵袭也减少。总之，这些数据首次证明了沙门氏菌针状复合体蛋白与肽聚糖之间存在关联。此外，我们证明这些蛋白质 - 肽聚糖相互作用对于这种特殊细胞器的高效和正确组装至关重要。
