Cornall, M. | Search bioRxiv

bioRxiv2024

ON-OFF nanopores for optical control of transmembrane ionic communication

[Abstract]Nanoscale photoswitchable proteins could facilitate precise spatiotemporal control of transmembrane communication and support studies in synthetic biology, neuroscience, and bioelectronics. Through covalent modification of the -hemolysin protein pore with arylazopyrazole photoswitches, we have produced "photopores" that transition between iontronic resistor and diode modes in response to irradiation at orthogonal wavelengths. In the diode mode, a low-leak OFF-state nanopore exhibits a reversible increase in unitary conductance of more than 20-fold upon irradiation at 365 nm. A rectification ratio of >5 was achieved with photopores in the diode state by either direct or alternating voltage input. Unlike conventional electronic phototransistors with intensity-dependent photoelectric responses, the photopores regulated current output solely based on the wavelength(s) of monochromatic or dual-wavelength irradiation. Dual-wavelength irradiation at various relative intensities allowed graded adjustment of photopore conductance. By using these properties, photonic signals were converted into ionic signals, highlighting the potential applications of photopores as components of smart devices in synthetic biology.

biophysics↗

bioRxiv2024

Multifunctional Systems in Synthetic Biology: Single-Stranded DNA Signaling for Precise Control of Gene Activation

Controllable gene circuits that respond to defined inputs are essential tools in synthetic biology. By leveraging regulatory mechanisms at either transcriptional or translational levels, synthetic responsive systems have been engineered to recognize diverse signals, such as small molecules (e.g., tetracycline) or physical stimuli (e.g., light). However, these approaches have limitations: small-molecule signals often require high concentrations to be effective, and sophisticated engineering is needed to generate responsive effectors. Here, we establish a simple, versatile gene activation system in which short single-stranded DNAs trigger RNA or protein production by complementing defective single-stranded promoters upstream of target genes. We demonstrate selective gene activation with orthogonal promoters, and logic-gate operations with signal pairs. The signaling system operates in compartmentalized nanoliter droplets scaffolded by bilayers. Signal delivery is controlled by selectively disrupting bilayers or applying transmembrane potential to move signals through protein pores, thereby activating genes within the receiver compartments. This work expands the toolset for engineering multifunctional, responsive materials to meet biotechnological and medical needs, enabling gene activation in response to specific cues.

synthetic biology↗