Identifying lipids tightly bound to an integral membrane protein

Area of Science:

• Membrane biophysics
• Microbial photosensors
• Biochemistry

Background:

• Anabaena Sensory Rhodopsin (ASR) from cyanobacteria is a microbial photosensor.
• Previous studies indicated ASR co-purifies with unknown small molecules.
• The structural and functional roles of these associated molecules were unclear.

Purpose of the Study:

• To characterize the small molecules associated with Anabaena Sensory Rhodopsin (ASR).
• To elucidate the structural and functional significance of these co-purifying molecules.
• To identify the specific sugars and lipids bound to ASR.

Main Methods:

• Solid-state nuclear magnetic resonance (SSNMR) spectroscopy.
• Mass spectrometry (MS).
• Phosphorus NMR and liquid chromatography-mass spectrometry (LC-MS).

Main Results:

• SSNMR spectra revealed patterns corresponding to Enterobacterial Common Antigen (ECA) sugars: N-acetyl-d-glucosamine, N-acetyl-d-mannosaminuronic acid, and 4-acetamido-4,6-dideoxy-d-galactose.
• Phosphorus NMR indicated monophosphates, suggesting phosphatidylglyceride as the ECA lipid carrier anchored to ASR.
• LC-MS confirmed the presence of phosphatidylethanolamine (PE) tightly interacting with ASR.

Conclusions:

• ASR tightly binds to ECA sugars and specific phospholipids, including phosphatidylglyceride and phosphatidylethanolamine.
• These findings provide insight into the molecular interactions and potential functional roles of ASR-associated molecules.
• This work honors Professor Michèle Auger's contributions to membrane biophysics and NMR.