Action spectrum and quantum efficiency for proton pumping in Halobacterium halobium

Area of Science:

• Microbiology
• Biophysics
• Photochemistry

Background:

• Halobacterium halobium utilizes bacteriorhodopsin for light-driven proton pumping.
• Proton influx can complicate measurements of proton ejection in intact cells.
• Carotenoids in H. halobium can absorb light, potentially interfering with action spectrum analysis.

Purpose of the Study:

• To determine the action spectrum and quantum efficiency for proton ejection in H. halobium.
• To investigate the role of carotenoids in light-driven proton transport.
• To develop a method for measuring bacteriorhodopsin activity in intact cells.

Main Methods:

• Measurement of action spectrum and quantum efficiency (phi H+) for proton ejection.
• Utilized conditions minimizing light-triggered proton influx.
• Developed a new nondestructive assay for bacteriorhodopsin in intact cells and envelopes.

Main Results:

• Carotenoid absorption does not contribute to proton ejection.
• The action spectrum, corrected for pigment shielding, matches bacteriorhodopsin's absorption spectrum.
• Quantum efficiency values (0.4-0.7) in cells and envelopes are higher than in isolated purple membrane.

Conclusions:

• Bacteriorhodopsin is the sole pigment responsible for light-induced proton ejection in H. halobium.
• The higher quantum efficiency suggests more than one proton is pumped per bacteriorhodopsin photocycle in intact cells and envelopes.
• The new assay allows for accurate assessment of bacteriorhodopsin function in complex biological samples.