Cell-Free Nonequilibrium Assembly for Hierarchical Protein/Peptide Nanopillars
Cells contain intricate protein nanostructures, but replicating them outside of cells presents challenges. One such example is the vertical fibronectin pillars observed in embryos. Here, we demonstrate the creation of cell-free vertical fibronectin pillar mimics using nonequilibrium self-assembly. Our approach utilizes enzyme-responsive phosphopeptides that assemble into nanotubes. Enzyme action triggers shape changes in peptide assemblies, driving the vertical growth of protein nanopillars into bundles. These bundles, with peptide nanotubes serving as a template to remodel fibronectin, can then recruit collagen, which forms aggregates or bundles depending on their types. Nanopillar formation relies on enzyme-catalyzed nonequilibrium self-assembly and is governed by the concentrations of enzyme, protein, peptide, the structure of the peptide, and peptide assembly morphologies. Cryo-EM reveals unexpected nanotube thinning and packing after dephosphorylation, indicating a complex sculpting process during assembly. Our study demonstrates a cell-free method for constructing intricate, multiprotein nanostructures with directionality and composition.
Authors
- Seth Fraden (ORCID: https://orcid.org/0000-0002-2420-9939)
- Junichi Usuba (ORCID: https://orcid.org/0000-0002-7571-5276)
- Grace G. D. Han (ORCID: https://orcid.org/0000-0002-2918-1584)
- Shane T. Rich-New (ORCID: https://orcid.org/0000-0002-2380-0778)
- Kangqiang Qiu (ORCID: https://orcid.org/0000-0002-8672-5648)
- Qian‐Feng Qiu (ORCID: https://orcid.org/0000-0002-4400-2142)
- Fengbin Wang (ORCID: https://orcid.org/0000-0003-1008-663X)
- Jiaqi Guo (ORCID: https://orcid.org/0000-0002-0254-0401)
- Meihui Yi (ORCID: https://orcid.org/0000-0002-1912-1043)
- Ayisha Zia (ORCID: https://orcid.org/0000-0002-9577-9584)
- Michael M. Norton (ORCID: https://orcid.org/0000-0002-1970-760X)
- Jiajie Diao (ORCID: https://orcid.org/0000-0003-4288-3203)
- Michael F. Hagan (ORCID: https://orcid.org/0000-0002-9211-2434)
- Bing Xu (ORCID: https://orcid.org/0000-0002-4639-387X)
- Zhiyu Liu (ORCID: https://orcid.org/0000-0001-9918-189X)
Institutions
- University of Alabama at Birmingham (US)
- University of Cincinnati (US)
- Brandeis University (US)
Publication Details
- Journal
- Journal of the American Chemical Society
- Published
- 2024-09-10
- DOI
- https://doi.org/10.1021/jacs.4c06775
- Citations
- 9
- Primary Topic
- Supramolecular Self-Assembly in Materials
- Type
- article
- Field-Weighted Citation Impact
- 1.56
Funders
- National Science Foundation
- Frederick National Laboratory for Cancer Research
- National Institutes of Health
- Materials Research Science and Engineering Center, Harvard University
- National Cancer Institute
- National Institute of General Medical Sciences
- Division of Materials Research