Temperature-resistant bicelles for structural studies by solid-state NMR spectroscopy

Area of Science:

• Biophysics
• Structural Biology
• Biochemistry

Background:

• Determining membrane protein structures is crucial for understanding biological functions.
• Challenges exist in maintaining native folding and function outside the cell membrane, especially for proteins with large soluble domains.
• Conventional micelles have limitations for structural studies, and while bicelles offer improvements, their thermal instability is a significant drawback.

Purpose of the Study:

• To overcome the temperature restrictions of bicelles for membrane protein structural studies.
• To identify optimal bicellar compositions for enhanced stability and broad temperature applicability.
• To demonstrate the utility of these temperature-resistant bicelles in solid-state NMR experiments.

Main Methods:

• Investigated bicelle compositions using 1,2-didecanoyl-sn-glycero-3-phosphocholine (DDPC) and 1,2-diheptanoyl-sn-glycero-3-phosphocholin (DHepPC).
• Assessed bicelle stability and magnetic alignment over a temperature range of -15 to 80 °C.
• Utilized two-dimensional separated-local field (SLF) solid-state NMR experiments on cytochrome b5 incorporated in aligned bicelles.
• Employed (31)P NMR experiments to characterize bicellar morphology, q ratio/size, and hydration levels.

Main Results:

• Identified DDPC and DHepPC bicelles as stable and resistant to temperature variations without additional stabilizers.
• Demonstrated robust bicellar phase and magnetic alignment across a broad temperature range (-15 to 80 °C).
• Showcased the retention of native membrane protein structure and increased sensitivity in low-temperature solid-state NMR experiments.
• Provided morphological information on DDPC-based bicelles, valuable for various biophysical and spectroscopic techniques.

Conclusions:

• Developed novel, temperature-resistant bicelles composed of DDPC and DHepPC that overcome limitations of conventional systems.
• These bicelles are suitable for structural studies of membrane proteins, including those with large soluble domains, across a wide temperature spectrum.
• The findings enhance the applicability of solid-state NMR, particularly at low temperatures, for membrane protein structure determination and biophysical investigations.