Enzyme- and DNAzyme-Driven Transient Assembly of DNA-Based Phase-Separated Coacervate Microdroplets

Area of Science:

• Synthetic biology
• Biochemistry
• Materials science

Background:

• Dynamic cellular processes rely on transient structures.
• DNA-based coacervates offer a platform for mimicking these dynamics.
• Controlling the assembly and disassembly of these structures is key for functional applications.

Purpose of the Study:

• To introduce enzyme- and DNAzyme-responsive DNA-based microdroplet (MD) coacervates.
• To demonstrate programmed and gated transient formation and depletion of MD frameworks.
• To explore MDs as functional carriers for temporal, dose-controlled release.

Main Methods:

• Utilized Y-shaped DNA frameworks cross-linked by complementary strands or palindromic strands.
• Engineered cleavage sites for endonucleases (EcoRI, HindIII) and nickases (Nt.BbvCI, Nb.BtsI).
• Incorporated Mg2+-DNAzyme units for metabolite-driven or light-modulated coacervate evolution and depletion.

Main Results:

• Demonstrated transient evolution and depletion of phase-separated MD coacervates using enzymes and DNAzymes.
• Achieved programmed and gated formation/depletion of MD frameworks by mixing responsive MDs.
• Showcased metabolite- and light-modulated dynamics of DNAzyme-functionalized MDs.

Conclusions:

• Enzyme- or DNAzyme-catalyzed transient MD coacervates provide protocell frameworks mimicking dynamic cellular processes.
• These systems offer a versatile platform for temporal, dose-controlled release applications.
• The study highlights the potential of DNA-based coacervates in synthetic biology and materials science.