Unfolding pathways of native bacteriorhodopsin depend on temperature

Area of Science:

• Biophysics
• Structural Biology
• Membrane Proteins

Background:

• Bacteriorhodopsin (BR) is a crucial membrane protein involved in light-driven proton pumping.
• Understanding the mechanical stability of BR's structural elements is vital for elucidating its function.
• Previous studies have explored BR structure, but detailed mechanical unfolding pathways at varying temperatures remain less understood.

Purpose of the Study:

• To investigate the mechanical stability of individual structural elements within bacteriorhodopsin.
• To determine the effect of physiologically relevant temperatures on the unfolding pathways of BR.
• To correlate temperature-induced changes in mechanical stability with alterations in unfolding mechanisms.

Main Methods:

• Utilized high-resolution atomic force microscopy (AFM) imaging.
• Employed single-molecule force spectroscopy to unfold single bacteriorhodopsins.
• Performed experiments on native purple membrane patches across a temperature range of 8 to 52 degrees C.

Main Results:

• Unfolding spectra provided detailed insights into the stability of alpha-helices and connecting loops.
• Unfolding forces of secondary structures significantly decreased with increasing temperature (8-52°C).
• Higher temperatures led to collective unfolding of transmembrane helices and loops, unlike individual unfolding at lower temperatures.

Conclusions:

• Temperature significantly influences the mechanical stability and unfolding pathways of bacteriorhodopsin.
• Elevated temperatures decrease the stability of secondary structural elements and alter inter-structure interactions.
• BR structural elements transition from individual unfolding barriers to collective unfolding at higher temperatures.