Measuring viscosity inside mesoporous silica using protein-bound molecular rotor probe

Area of Science:

• Materials Science
• Biophysics
• Nanotechnology

Background:

• Mesoporous silica materials (SBA-15 and MCF) are widely used for enzyme immobilization.
• Understanding the microenvironment within these pores is crucial for optimizing protein behavior and activity.
• Fluorescence spectroscopy offers a sensitive method to probe local conditions.

Purpose of the Study:

• To measure the effective viscosity inside mesoporous silica pores using protein-bound molecular rotors.
• To investigate the influence of protein concentration and pore surface modification on viscosity and protein distribution.
• To assess the role of Förster Resonance Energy Transfer (FRET) in fluorescence-based measurements within confined environments.

Main Methods:

• Utilizing fluorescence spectroscopy of cyanine dyes (Cy3 and Cy5) as molecular rotors to probe viscosity.
• Calibrating dye response using glycerol/water mixtures and applying the intensity ratio method.
• Immobilizing lipase onto SBA-15 and MCF silica particles, with some MCF particles modified with octyl groups.
• Analyzing FRET between dyes attached to the same or different proteins to understand protein proximity and distribution.

Main Results:

• Effective viscosity inside mesoporous silica pores is one order of magnitude higher than in bulk water.
• Protein-protein interactions are significantly stronger within the pores compared to bulk solutions.
• Octyl-modified MCF particles showed enhanced lipase uptake and FRET between dyes on different proteins, indicating non-homogenous protein distribution.
• Dye emission redshift suggests a less polar pore environment in modified particles, correlating with higher immobilization efficiency.

Conclusions:

• Mesoporous silica pores create a high-viscosity environment that influences protein behavior.
• Surface modification of mesoporous silica can alter protein distribution and interactions within pores.
• Fluorescence methods, including FRET, provide valuable insights into protein immobilization and pore microenvironments.
• Understanding these factors is key to optimizing enzyme immobilization for biotechnological applications.