ArticleScientific reports2022
Cell-free protein crystallization for nanocrystal structure determination.
Article in Scientific reports, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 8 papers.
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Who cites it
8 citing papers in PubMed.
- A note on the appearance of PEG in macromolecular crystals.Acta crystallographica. Section F, Structural biology communications · 2026Article
- Half a Century of Progress: The Evolution of Wheat Germ-Based In Vitro Translation into a Versatile Protein Production Method.International journal of molecular sciences · 2025Review
- Macromolecular crystallography at SPring-8 and SACLA.Journal of synchrotron radiation · 2025Article
- Useful experimental aspects of small-wedge synchrotron crystallography for accurate structure analysis of protein molecules.Acta crystallographica. Section D, Structural biology · 2025Article
- Review
- High-throughput structure determination of an intrinsically disordered protein using cell-free protein crystallization.Proceedings of the National Academy of Sciences of the United States of America · 2024Article
- Efficiency of transcription and translation of cell-free protein synthesis systems in cell-sized lipid vesicles with changing lipid composition determined by fluorescence measurements.Scientific reports · 2024Article
- Solid-Phase Cell-Free Protein Synthesis and Its Applications in Biotechnology.Advances in biochemical engineering/biotechnology · 2023Article
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Authors and funding
8 authors.
Funding
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Abstract
In-cell protein crystallization (ICPC) has been investigated as a technique to support the advancement of structural biology because it does not require protein purification and a complicated crystallization process. However, only a few protein structures have been reported because these crystals formed incidentally in living cells and are insufficient in size and quality for structure analysis. Here, we have developed a cell-free protein crystallization (CFPC) method, which involves direct protein crystallization using cell-free protein synthesis. We have succeeded in crystallization and structure determination of nano-sized polyhedra crystal (PhC) at a high resolution of 1.80 Å. Furthermore, nanocrystals were synthesized at a reaction scale of only 20 μL using the dialysis method, enabling structural analysis at a resolution of 1.95 Å. To further demonstrate the potential of CFPC, we attempted to determine the structure of crystalline inclusion protein A (CipA), whose structure had not yet been determined. We added chemical reagents as a twinning inhibitor to the CFPC solution, which enabled us to determine the structure of CipA at 2.11 Å resolution. This technology greatly expands the high-throughput structure determination method of unstable, low-yield, fusion, and substrate-biding proteins that have been difficult to analyze with conventional methods.
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