Monitoring physical and chemical properties of individual carboxysomes trapped in solution

Abstract

Biological nano-objects rapidly diffuse in the solution phase, impeding our efforts to monitor their physical and chemical properties for extended periods of time. To overcome this, we developed the Interferometric Scattering Anti-Brownian ELectrokinetic (ISABEL) Trap, which counteracts Brownian motion for an extended time by tracking a particle's location via its scattering and rapidly applying positional feedback with electrokinetic forces. Recently, we improved the flexibility of these experiments by shifting the scattering detection beam to the near-infrared and opening the visible region for flexible and specific fluorescence measurements. These capabilities allow us to monitor the physical and chemical properties of the carboxysome, a ~100nm bacterial microcompartment responsible for CO2 fixation by the enzyme Rubisco. With the ISABEL trap, we can rapidly interleave 405 and 488 nm fluorescence excitation beams to measure the redox properties inside individual carboxysomes using the redox reporting GFP mutant, roGFP2. The capabilities provided by the ISABEL trap allow us to design solution-phase single-particle experiments for a variety of biological nanoscale objects.