Decoding the absolute stoichiometric composition and structural plasticity of α-carboxysomes

Carboxysomes are anabolic bacterial microcompartments that play an essential role in carbon fixation in cyanobacteria and some chemoautotrophs. This self-assembling organelle encapsulates the key CO2-fixing enzymes, Rubisco, and carbonic anhydrase using a polyhedral protein shell that is constructed by hundreds of shell protein paralogs. The α-carboxysome from the chemoautotroph Halothiobacillus neapolitanus serves as a model system in fundamental studies and synthetic engineering of carboxysomes. Here we adopt a QconCAT-based quantitative mass spectrometry approach to determine the stoichiometric composition of native α-carboxysomes from H. neapolitanus. We further performed an in-depth comparison of the protein stoichiometry of native and recombinant α-carboxysomes heterologously generated in Escherichia coli to evaluate the structural variability and remodeling of α-carboxysomes. Our results provide insight into the molecular principles that mediate carboxysome assembly, which may aid in rational design and reprogramming of carboxysomes in new contexts for biotechnological applications

羧酶体（Carboxysomes）是一类同化性细菌微区室，在蓝细菌与部分化能自养菌的碳固定过程中发挥不可或缺的关键作用。该自组装细胞器的多面体形蛋白外壳由数百种壳蛋白旁系同源蛋白构建，用以包裹核心的CO₂固定酶——核酮糖-1,5-二磷酸羧化酶/加氧酶（Rubisco）与碳酸酐酶。来自化能自养菌那不勒斯嗜硫杆菌（Halothiobacillus neapolitanus）的α-羧酶体，是羧酶体基础研究与合成工程领域的经典模型系统。本研究采用基于QconCAT的定量质谱分析方法，测定了来自那不勒斯嗜硫杆菌的天然α-羧酶体的化学计量组成。我们进一步对天然α-羧酶体与在大肠杆菌（Escherichia coli）中异源表达生成的重组α-羧酶体的蛋白化学计量进行了深入比较，以评估α-羧酶体的结构变异与重塑过程。本研究结果揭示了介导羧酶体组装的分子原理，可为在生物技术应用的全新场景下理性设计与重编程羧酶体提供助力。