Scheffel André, Schüler Dirk
Max Planck Institute for Marine Microbiology, Bremen, Germany.
J Bacteriol. 2007 Sep;189(17):6437-46. doi: 10.1128/JB.00421-07. Epub 2007 Jun 29.
Magnetotactic bacteria navigate along the earth's magnetic field using chains of magnetosomes, which are intracellular organelles comprising membrane-enclosed magnetite crystals. The assembly of highly ordered magnetosome chains is under genetic control and involves several specific proteins. Based on genetic and cryo-electron tomography studies, a model was recently proposed in which the acidic MamJ magnetosome protein attaches magnetosome vesicles to the actin-like cytoskeletal filament formed by MamK, thereby preventing magnetosome chains from collapsing. However, the exact functions as well as the mode of interaction between MamK and MamJ are unknown. Here, we demonstrate that several functional MamJ variants from Magnetospirillum gryphiswaldense and other magnetotactic bacteria share an acidic and repetitive central domain, which displays an unusual intra- and interspecies sequence polymorphism, probably caused by homologous recombination between identical copies of Glu- and Pro-rich repeats. Surprisingly, mamJ mutant alleles in which the central domain was deleted retained their potential to restore chain formation in a DeltamamJ mutant, suggesting that the acidic domain is not essential for MamJ's function. Results of two-hybrid experiments indicate that MamJ physically interacts with MamK, and two distinct sequence regions within MamJ were shown to be involved in binding to MamK. Mutant variants of MamJ lacking either of the binding domains were unable to functionally complement the DeltamamJ mutant. In addition, two-hybrid experiments suggest both MamK-binding domains of MamJ confer oligomerization of MamJ. In summary, our data reveal domains required for the functions of the MamJ protein in chain assembly and maintenance and provide the first experimental indications for a direct interaction between MamJ and the cytoskeletal filament protein MamK.
趋磁细菌利用磁小体链沿着地球磁场导航，磁小体是由膜包裹的磁铁矿晶体组成的细胞内细胞器。高度有序的磁小体链的组装受基因控制，涉及几种特定蛋白质。基于遗传学和冷冻电子断层扫描研究，最近提出了一个模型，其中酸性的MamJ磁小体蛋白将磁小体囊泡附着到由MamK形成的肌动蛋白样细胞骨架丝上，从而防止磁小体链坍塌。然而，MamK和MamJ之间的确切功能以及相互作用模式尚不清楚。在这里，我们证明来自嗜格氏嗜磁螺菌和其他趋磁细菌的几种功能性MamJ变体共享一个酸性且重复的中央结构域，该结构域显示出不寻常的种内和种间序列多态性，可能是由富含Glu和Pro的重复序列的相同拷贝之间的同源重组引起的。令人惊讶的是，缺失中央结构域的mamJ突变等位基因在ΔmamJ突变体中仍保留恢复链形成的潜力，这表明酸性结构域对于MamJ的功能不是必需的。双杂交实验结果表明MamJ与MamK发生物理相互作用，并且MamJ内两个不同的序列区域被证明参与与MamK的结合。缺乏任何一个结合结构域的MamJ突变变体无法在功能上互补ΔmamJ突变体。此外，双杂交实验表明MamJ的两个MamK结合结构域都赋予MamJ寡聚化。总之，我们的数据揭示了MamJ蛋白在链组装和维持功能中所需的结构域，并为MamJ与细胞骨架丝蛋白MamK之间的直接相互作用提供了首个实验证据。
