The assembly of protein machines in cells is precise, rapid, and coupled to protein synthesis with regulation in space and time. The assembly of natural and synthetic nanomachines could be similarly controlled by genetic programming outside the cell. Here, we present quasi-two-dimensional (2D) silicon compartments that enable programming of protein assembly lines by local synthesis from surface-immobilized DNA brushes. Using this platform, we studied the autonomous synthesis and assembly of a structural complex from a bacteriophage and a bacterial RNA-synthesizing machine. Local synthesis and surface capture of complexes provided high assembly yield and sensitive detection of spatially resolved assembly intermediates, with the 3D geometry of the compartment and the 2D pattern of brushes dictating the yield and mode of assembly steps. Localized synthesis of proteins in a single gene brush enhances their interactions, and displacement of their genes in separated brushes leads to step-by-step surface assembly. This methodology enables spatial regulation of protein synthesis, and deciphering, reconstruction and design of biological machine assembly lines.
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The data necessary to interpret, replicate and build upon this work appears in the article and its Supplementary information. Additional data can be made available from the corresponding authors upon reasonable request.
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We acknowledge funding from the Israel Science Foundation (grant no. 1870/15), the United States–Israel Binational Science Foundation (grant no. 2014400), and the Minerva Foundation (grant no. 712274) for the work on the T4 wedges. We thank the Office of Naval Research (award no. N62909-18-1-2094) for funding the work on RNAP assembly. We thank M. Levy for discussions.
The authors declare no competing interests.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
a, Scheme, co-expression and interaction of gp10 and gp7 in solution prior to surface binding sequester the scaffolding of gp7 onto gp10 pre-bound to the surface. b, Dose response of gene-10 fraction in a mixed DNA brush with fixed amount of gene-7. Surface gp11 traps were pre-bound with gp10. Surface bound gp10-7 complexes revealed by post-staining with FL-gp8. Detailed gene composition of all experiments appears in Supplementary Table 3. Individual data points and mean values are shown, error bars represent ± s.d. The number of samples for each data point (b) is listed in Supplementary Table 4.
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Vonshak, O., Divon, Y., Förste, S. et al. Programming multi-protein assembly by gene-brush patterns and two-dimensional compartment geometry. Nat. Nanotechnol. 15, 783–791 (2020). https://doi.org/10.1038/s41565-020-0720-7