Measurement of Macromolecular Crowding in Rhodobacter sphaeroides under Different Growth Conditions

Area of Science:

• Microbiology
• Cell Biology
• Biophysics

Background:

• Bacterial cytoplasm is a crowded environment.
• Cytoplasmic crowding influences protein folding, diffusion, and complex formation.
• Previous studies often imposed stress to alter crowding.

Purpose of the Study:

• Investigate the effects of naturally induced cytoplasmic crowding in Rhodobacter sphaeroides.
• Determine how changes in intracytoplasmic membrane (ICM) vesicles affect protein diffusion and conformation.

Main Methods:

• Utilized a fluorescence resonance energy transfer (FRET) sensor to measure protein conformation.
• Employed photoactivated localization microscopy (PALM) to assess protein diffusion coefficients.
• Cultured Rhodobacter sphaeroides under varying light intensities to modulate ICM vesicle numbers.

Main Results:

• Low light conditions (high ICM vesicles) led to a more condensed FRET probe conformation.
• Protein diffusion coefficients for molecules >27 kDa decreased with increasing ICM vesicle numbers.
• Protein diffusion was slower in low light (high crowding) compared to high light/aerobic conditions.

Conclusions:

• Naturally altering cytoplasmic crowding via ICM vesicles affects protein conformation and diffusion rates in Rhodobacter sphaeroides.
• Changes in cytoplasmic crowding can influence the kinetics of physiological activities.
• Findings have implications for bacterial growth and responses under dynamic environmental conditions.