A Quasi-2D Platform for Programming and Detecting Macromolecular Self-Assembly
Oral-Virtual · Withdrawn
Abstract
The precise and rapid assembly of multi-protein machines within biological cells is tightly regulated in both space and time, and closely coupled to protein synthesis. A similar process should be achievable outside of the living cell. Natural and synthetic nanomachines could be synthesized and assembled on synthetic platforms, following genetic programming.
Here, we present a quasi-2D on-chip platform enabling the programmable synthesis and self-assembly of multi-protein machines. This platform is based on local protein synthesis from dense surface-immobilized DNA brushes. The confinement to the surface creates the necessary biophysical conditions at the nanoscale for concomitant synthesis and interactions of parts, and for the visualization of nascent multi-protein assemblies.
Using this platform, we successfully reconstituted the autonomous synthesis and assembly of the E. coli’s small ribosomal subunit, accomplished by expressing one ribosomal RNA, twenty ribosomal proteins, and six assembly factors from the surface-immobilized genes. Our real-time detection showed hierarchal binding, cooperative interactions, unstable intermediates, and specific binding to large ribosomal subunits. This platform also promoted the self-assembly of late-stage intermediates of the large subunit.
Finally, the system was extended toward more cell-like conditions, integrating lipid-bound compartmentalization with DNA brushes.
Here, we present a quasi-2D on-chip platform enabling the programmable synthesis and self-assembly of multi-protein machines. This platform is based on local protein synthesis from dense surface-immobilized DNA brushes. The confinement to the surface creates the necessary biophysical conditions at the nanoscale for concomitant synthesis and interactions of parts, and for the visualization of nascent multi-protein assemblies.
Using this platform, we successfully reconstituted the autonomous synthesis and assembly of the E. coli’s small ribosomal subunit, accomplished by expressing one ribosomal RNA, twenty ribosomal proteins, and six assembly factors from the surface-immobilized genes. Our real-time detection showed hierarchal binding, cooperative interactions, unstable intermediates, and specific binding to large ribosomal subunits. This platform also promoted the self-assembly of late-stage intermediates of the large subunit.
Finally, the system was extended toward more cell-like conditions, integrating lipid-bound compartmentalization with DNA brushes.
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Publication: [1] M. Levy et al, Autonomous synthesis and assembly of a ribosomal subunit on a chip, Sci. Adv. 2020 6, eaaz6020.
[2] M. Levy et al, Boundary-free ribosome compartmentalization by gene expression on a surface, ACS Synth. Biol., 2021 10, 3, 609–619
[3] N. Avidan et al, Toward Memory in a DNA Brush: Site-Specific Recombination Responsive to Polymer Density, Orientation, and Conformation, J. Am. Chem. Soc. 2023, 145, 17, 9729–9736
Presenters
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Michael Levy
- Technion - Israel Institute of Technology