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Observations of phase changes in monoolein during high viscous injection.
Daniel J Wells1, Peter Berntsen1, Eugeniu Balaur1
1La Trobe Institute for Molecular Science, Department of Mathematical and Physical Sciences, School of Computing Engineering and Mathematical Science, La Trobe University, Bundoora, VIC 3086, Australia.
Journal of Synchrotron Radiation
|May 5, 2022
Summary
High-viscosity injectors (HVIs) can alter lipidic cubic phases (LCPs) used for membrane protein crystallography. This study reveals LCP phase changes during injection, impacting diffraction data quality.
Area of Science:
- Structural biology
- Biophysics
- Crystallography
Background:
- Membrane protein crystallography often uses lipidic cubic phases (LCPs) as a carrier medium delivered via high-viscosity injectors (HVIs).
- The structural integrity of LCPs during the injection process is crucial for obtaining reliable X-ray diffraction data but remains poorly understood.
- Potential structural alterations in LCPs due to pressure, dehydration, and temperature changes during injection could affect experimental outcomes.
Purpose of the Study:
- To investigate the structural and phase behavior of monoolein/water and monoolein/buffer mixtures within HVIs.
- To determine the influence of continuous flow injection, pressure variations, and vacuum conditions on LCP structure.
- To correlate HVI reservoir pressure with observed LCP phase changes and lattice parameter shifts.
Main Methods:
- Studied phase changes in monoolein/water and monoolein/buffer mixtures during continuous flow injection under atmospheric and vacuum conditions.
- Monitored HVI reservoir pressure to assess its correlation with LCP phase behavior.
- Employed optical polarization microscopy and simulation studies to analyze LCP structural changes.
Main Results:
- HVI reservoir pressure variations did not consistently correlate with observed LCP phase changes or lattice parameter shifts.
- Vacuum injection induced a three-way phase coexistence: gyroid cubic, diamond cubic, and lamellar phases.
- Atmospheric pressure injection showed coexistence of cubic and lamellar phases, with lamellar phase formation dependent on co-flowing gas conditions.
Conclusions:
- The injection process significantly impacts LCP structure, leading to phase transitions that can affect membrane protein crystal diffraction data.
- Understanding and controlling these LCP phase changes under HVI conditions is essential for optimizing serial crystallography experiments.
- Co-flowing gas conditions play a critical role in stabilizing the LCP stream and mitigating unwanted lamellar phase formation.
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