Boundary-Free Ribosome Compartmentalization by Gene Expression on a Surface

Area of Science:

• Synthetic biology
• Biophysics
• Molecular systems engineering

Background:

• Artificial cell models aim to replicate the compartmentalization found in living cells.
• Dense DNA brushes act as synthetic operons for autonomous synthesis and assembly of cellular machinery.

Purpose of the Study:

• To investigate ribosome compartmentalization within minimal gene-expression reactions on a surface.
• To explore transcription-translation coupling and protein synthesis in engineered surface-bound compartments.

Main Methods:

• Utilized evanescent field fluorescence microscopy to observe molecular colocalization within DNA brushes.
• Employed fluorescence recovery after photobleaching to analyze ribosome diffusion dynamics.
• Developed surface-based systems with immobilized ribosomes adjacent to DNA brushes to decouple transcription and translation.

Main Results:

• Observed colocalization of transcription and translation machinery within dense DNA brushes, indicating coupled gene expression.
• Demonstrated significantly reduced ribosome diffusion and increased local concentration within DNA brushes, forming a functional compartment.
• Showcased the ability of immobilized ribosomes to synthesize proteins, creating 2D subcompartments regulated by DNA brush design.

Conclusions:

• Dense DNA brushes can create boundary-free, ribosome-rich compartments for coupled transcription-translation.
• Decoupling transcription and translation by immobilizing ribosomes allows for controlled protein synthesis and patterned subcompartments.
• Further surface-based localization of molecular components can enhance compartmentalization of gene-expression reactions.