Enzyme Fueled Dissipative Self-assembly of Guanine Functionalized Molecules and Their Cellular Behaviour

Generally, an esterase lipase enzyme can hydrolyze specific substrates called esters in an aqueous solution. Herein, we investigate how a G-quadruplex self-assembly affects the hydrolysis equilibrium in reverse. The biocatalyst, lipase, activates the individual building-blocks through fuel consumption, causing them to undergo a higher degree of self-organization into nanofibers within spheres. We have synthesized five peptide-lipid-conjugated guanine base functionalized molecules to explore how the equilibrium can be shifted through reverse hydrolysis. Among these, NAC5 self-assembled into a G-quadruplex structure which has been confirmed by various spectroscopic techniques. The wide-angle powder XRD, ThT dye binding assay and circular dichroism study is carried out to support the presence of the G-quadruplex structure. The biocatalytic formation of nanofibers enclosed spheres is analyzed using CLSM, FE-SEM and HR-TEM experiments. Additionally, we assess the biocompatibility of the enzyme fueled dissipative self-assembled fibers enclosed spheres, as they have potential applications as a biomaterial in protocells. MTT assay is performed to check the cytotoxicity of G-quadruplex hydrogel, using HEK 293 and McCoy cell lines for viability assessment. Finally, the utility of the novel NAC5 hydrogel as a wound repairing biomaterial is demonstrated by cell migration experiment in a scratch assay.