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Microfluidic Mixers for Studying Protein Folding
Published on: April 10, 2012
Nonequilibrium single molecule protein folding in a coaxial mixer
Kambiz M Hamadani1, Shimon Weiss
1Department of Chemistry and Biochemistry, University of California at Los Angeles, Los Angeles, California, USA. khamadan@chem.ucla.edu
Biophysical Journal
|March 15, 2008
Summary
Researchers created a rapid, capillary-based mixer for single-molecule biophysics. This device precisely tracks protein folding dynamics, revealing details about the unfolded state of Chymotrypsin Inhibitor Protein 2.
Area of Science:
- Biophysics
- Chemical Engineering
- Materials Science
Background:
- Single-molecule biophysics requires precise control over reaction conditions.
- Existing microfluidic mixers can be expensive and complex to fabricate.
- Rapid mixing is crucial for studying fast biological processes.
Purpose of the Study:
- To develop and characterize a novel continuous-flow mixing device for single-molecule bioconformational studies.
- To enable rapid, in-lab construction of microfluidic mixers.
- To apply the device to investigate protein folding dynamics.
Main Methods:
- Continuous-flow mixing device with coaxial geometry for hydrodynamic focusing.
- Capillary-based design for ease of construction.
- Fluorescence cross-correlation spectroscopy velocimetry and finite element fluid dynamics simulations for characterization.
- Single-molecule Förster Resonance Energy Transfer (smFRET) for protein folding studies.
Main Results:
- The device achieves a response time of approximately 10 ms.
- Hydrodynamic focusing ensures rapid and efficient diffusional mixing.
- In-line filtering extends device lifetime to many months.
- Successfully resolved denaturant-dependent collapse from barrier-limited folding of Chymotrypsin Inhibitor Protein 2 (CI2).
Conclusions:
- The developed mixer is a cost-effective and efficient tool for single-molecule biophysics.
- It allows for high-resolution temporal studies of protein folding.
- The findings support the existence of a heterogeneous unfolded state in CI2, including cis- and trans-proline conformers.
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