Breindel Leonard, DeMott Christopher, Burz David S, Shekhtman Alexander
Department of Chemistry, University at Albany, State University of New York , 1400 Washington Avenue, Albany, New York 12222, United States.
Biochemistry. 2018 Feb 6;57(5):540-546. doi: 10.1021/acs.biochem.7b00938. Epub 2018 Jan 8.
How ribosome antibiotics affect a wide range of biochemical pathways is not well understood; changes in RNA-mediated protein quinary interactions and consequent activity inside the crowded cytosol may provide one possible mechanism. We developed real-time (RT) in-cell nuclear magnetic resonance (NMR) spectroscopy to monitor temporal changes in protein quinary structure, for ≥24 h, in response to external and internal stimuli. RT in-cell NMR consists of a bioreactor containing gel-encapsulated cells inside a 5 mm NMR tube, a gravity siphon for continuous exchange of medium, and a horizontal drip irrigation system to supply nutrients to the cells during the experiment. We showed that adding antibiotics that bind to the small ribosomal subunit results in more extensive quinary interactions between thioredoxin and mRNA. The results substantiate the idea that RNA-mediated modulation of quinary protein interactions may provide the physical basis for ribosome inhibition and other regulatory pathways.
核糖体抗生素如何影响广泛的生化途径尚不清楚；RNA介导的蛋白质五元相互作用的变化以及在拥挤的细胞质中随之而来的活性变化可能提供一种可能的机制。我们开发了实时（RT）细胞内核磁共振（NMR）光谱技术，以监测蛋白质五元结构在≥24小时内响应外部和内部刺激的时间变化。RT细胞内NMR由一个生物反应器组成，该生物反应器在一个5毫米NMR管内包含凝胶包裹的细胞、一个用于连续更换培养基的重力虹吸管以及一个在实验期间为细胞供应营养的水平滴灌系统。我们发现，添加与小核糖体亚基结合的抗生素会导致硫氧还蛋白和mRNA之间更广泛的五元相互作用。这些结果证实了RNA介导的五元蛋白质相互作用调节可能为核糖体抑制和其他调节途径提供物理基础的观点。
