Semi-Independent Control of Stability and Mobility in DNA Condensates

Liquid-like biomolecular condensates possess unique physical properties that are essential for cellular functions and artificial cell engineering. However, increasing the thermal stability of these condensates typically reduces their liquid-like dynamics. Here, we report a strategy to semi-independently control the thermal stability and dynamic properties of DNA droplets formed from six-branched DNA nanostructures (S-motifs). We designed S-motifs containing four fixed sticky ends (SEs) composed of 4 nucleotides (nt) and two variable-length SEs of 0-20 nt in length. Extending the length of the two variable SEs increased the phase-separation temperature (T_p) of the droplets. Within a specific range (4-12 nt), where the melting temperature (T_m) of the variable SEs remained below T_p, the droplets maintained their fusion dynamics and internal mobility despite increased thermal stability. In contrast, when the variable SE length was 16-20 nt and T_m exceeded T_p, the dynamic behaviors were inhibited because of stable polymerization. These findings demonstrate that the partial modification of the SE binding strength enables the tuning of thermal stability without sacrificing liquid fluidity, providing a valuable design principle for developing functional DNA-based artificial cells and molecular robots.