Proteorhodopsin: characterisation of 2D crystals by electron microscopy and solid state NMR

Area of Science:

• Biochemistry
• Structural Biology
• Microbiology

Background:

• Proteorhodopsin (PR) is a light-activated proton pump found in gamma-proteobacteria.
• It utilizes light energy for cellular energy production, analogous to bacteriorhodopsin.
• Numerous PR variants exist, spectrally tuned to marine light conditions.

Purpose of the Study:

• To characterize the structure of a green proteorhodopsin variant using 2D crystallization.
• To investigate the oligomeric state and protein packing of PR.

Main Methods:

• Electron microscopy (EM) for 2D crystal imaging.
• Solid-state Nuclear Magnetic Resonance (NMR) spectroscopy (15N MAS NMR, Lee-Goldberg cross-polarization) for structural and mobility analysis.
• Preparation of isotopically labeled PR samples (lysine, tryptophan, methionine).

Main Results:

• Stable 2D crystals of green PR with hexagonal packing were formed under diverse conditions.
• A low-resolution EM map showed PR forming ring-shaped oligomers.
• Solid-state NMR confirmed the protonated Schiff base chemical shift and probed backbone mobility.

Conclusions:

• Proteorhodopsin likely forms closely packed oligomeric structures in its native state.
• The study provides structural insights into PR's function as a proton pump.
• Characterization of PR structure aids in understanding microbial energy transduction in marine ecosystems.