National Institutes of Health/National Institute of Diabetes and Digestive and Kidney Disease (NIH/NIDDK)
ZIA-DK075136
United States
Citation
Journal: Nat Struct Mol Biol / Year: 2019 Title: Structural basis of amino acid surveillance by higher-order tRNA-mRNA interactions. Authors: Shuang Li / Zhaoming Su / Jean Lehmann / Vassiliki Stamatopoulou / Nikoleta Giarimoglou / Frances E Henderson / Lixin Fan / Grigore D Pintilie / Kaiming Zhang / Muyuan Chen / Steven J Ludtke ...Authors: Shuang Li / Zhaoming Su / Jean Lehmann / Vassiliki Stamatopoulou / Nikoleta Giarimoglou / Frances E Henderson / Lixin Fan / Grigore D Pintilie / Kaiming Zhang / Muyuan Chen / Steven J Ludtke / Yun-Xing Wang / Constantinos Stathopoulos / Wah Chiu / Jinwei Zhang / Abstract: Amino acid availability in Gram-positive bacteria is monitored by T-box riboswitches. T-boxes directly bind tRNAs, assess their aminoacylation state, and regulate the transcription or translation of ...Amino acid availability in Gram-positive bacteria is monitored by T-box riboswitches. T-boxes directly bind tRNAs, assess their aminoacylation state, and regulate the transcription or translation of downstream genes to maintain nutritional homeostasis. Here, we report cocrystal and cryo-EM structures of Geobacillus kaustophilus and Bacillus subtilis T-box-tRNA complexes, detailing their multivalent, exquisitely selective interactions. The T-box forms a U-shaped molecular vise that clamps the tRNA, captures its 3' end using an elaborate 'discriminator' structure, and interrogates its aminoacylation state using a steric filter fashioned from a wobble base pair. In the absence of aminoacylation, T-boxes clutch tRNAs and form a continuously stacked central spine, permitting transcriptional readthrough or translation initiation. A modeled aminoacyl disrupts tRNA-T-box stacking, severing the central spine and blocking gene expression. Our data establish a universal mechanism of amino acid sensing on tRNAs and gene regulation by T-box riboswitches and exemplify how higher-order RNA-RNA interactions achieve multivalency and specificity.
Mass: 18.015 Da / Num. of mol.: 18 / Source method: isolated from a natural source / Formula: H2O
Has ligand of interest
N
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Experimental details
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Experiment
Experiment
Method: X-RAY DIFFRACTION / Number of used crystals: 1
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Sample preparation
Crystal
Density Matthews: 2.52 Å3/Da / Density % sol: 51.18 % Description: Rectangular prism-shaped crystals grew in 2-7 days to maximum dimensions of 300 x 300 x 50 micrometer3.
Crystal grow
Temperature: 293 K / Method: vapor diffusion, hanging drop / pH: 7.4 Details: 0.1 M Morpheus Buffer System 2 pH 7.5 (Sodium HEPES; MOPS (acid)), 0.09 M Halogens (NaF; NaBr; NaI), 10% w/v PEG 20000, and 20% v/v PEG 500 monomethyl ether (MME)
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Data collection
Diffraction
Mean temperature: 100 K / Serial crystal experiment: N
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