OxyR调控磁螺菌MSR-1磁小体结构和组分变化的分子机理

Magnetotactic bacteria, a special kind of bacteria, synthesize intracellular nano-scale (40-100 nm) magnetic particles (magnetosomes). These bacteria have attracted much attention as model organisms to understand the mechanism and physiological significance of the synthesis of magnetic particles in higher organisms, and because of broad application prospects of these particles. However, molecular mechanisms and physiological functions of magnetosomes synthesis have not yet been understood. Several hypotheses have been proposed, which speculated the existence of an intermediate state of Fe2O3 • nH2O compounds during Fe3O4 synthesis process, but no direct evidence. Our previous work for the first time indicated peroxidase-like catalytic properties of magnetosomes and one of the physiological functions of these particles is the scavenging of intracellular reactive oxygen species (ROS) to protect cells against oxidative stress. The gene mutation experiments prove that OxyR disruptation mutant loss magnetic and magnetosomes crystals are smaller than that of wild type. Preliminary experiments of crystal diffraction analysis of magnetosomes in mutant under high-resolution electron microscopy indicate that it is composed of multiple iron oxides. To obtain conclusive evidence of key roles of OxyR which control changes of magnetosome from Fe2O3 to Fe3O4, and clear the regulatory molecular mechanism and model, we present the application of this project. It will provide a direct evidence for Fe2O3 • nH2O intermediate state compounds during magnetosome synthesis process and perfect the previous molecular models of magnetosomes formation.

趋磁细菌是一类合成40-100nm、膜包被的Fe3O4（磁小体）的特殊细菌，是研究高等生物合成磁性颗粒机理和生理意义的模式生物。同时磁小体具有广泛的应用前景。趋磁细菌磁小体合成的分子机制和生理功能至今尚未明了，多个假设模型已被提出，其中都推测Fe3O4合成过程中存在Fe2O3•nH2O中间态化合物，但无直接证据。我们的前期工作首次明确了磁小体具有类似过氧化物酶的催化性质，生理功能之一是清除氧化胁迫产生的自由基以保护细胞自身。抗氧化胁迫转录因子OxyR突变株的磁性消失、磁小体晶体变小，高分辨电子显微镜单胞晶体衍射结构和组分分析证明突变株中存在多种铁氧化合物的磁小体。为此提出本课题，拟确证OxyR调控磁小体由不同Fe2O3向Fe3O4转变的关键功能并明确调控的分子机理，建立调控模型。补充并丰富已有的磁小体合成分子模型，为磁小体合成过程中Fe2O3•nH2O中间态化合物的存在提供直接证据。

趋磁细菌(magnetotactic bacteria，MTB)的典型特征是在胞内合成纳米磁小体(magnetosomes，成分为Fe3O4，大小为40-50nm)，且可以沿地球磁力线方向运动。磁小体可作为酶、抗原、抗体、基因以及药物的载体，而广泛的应用于固定化酶、高灵敏度的免疫检测、基因治疗及肿瘤的靶向治疗等，但迄今为止其合成的分子机理和生理意义尚不清楚。本课题通过突变OxyR-like，比较野生型、突变体及互补菌株表型揭示OxyR-like控制磁小体的合成。高分辨电镜分析突变体和野生型胞内合成的单胞晶体组分证明 epsilon -Fe2O3和 a-Fe2O3是磁小体合成的中间体，其中 epsilon-Fe2O3是首次发现的中间体。凝胶阻滞和qRT-PCR实验证明OxyR-like调控自身及丙酮酸脱氢酶基因簇的表达，当OxyR-Like缺失时，细胞中三羧酸循环受到严重抑制，导致碳代谢速度减慢，能量和还原力供给不足。结论：OxyR-like通过调控碳代谢而控制磁小体合成。生物信息学分析发现胞内还有另一个与OxyR-like高度同源的OxyR（MGMSRv2_4250），其功能分析正在进行中，初步研究结果证明其为全局转录调控因子，通过调控胞内氧化还原状态等调控磁小体的合成。