Solutes modify a conformational transition in a membrane transport protein

Area of Science:

• Biochemistry
• Structural Biology
• Membrane Transport

Background:

• The bacterial outer-membrane vitamin B(12) transporter, BtuB, exhibits a significant order-to-disorder transition in its N-terminal energy-coupling motif (Ton box) upon substrate binding.
• This conformational change is crucial for the transporter's function.

Purpose of the Study:

• To investigate the effect of various solutes on the BtuB transporter's conformational transition.
• To elucidate the mechanism by which solutes influence the Ton box structure.

Main Methods:

• Site-directed spin labeling (SDSL) was employed to monitor conformational changes in BtuB.
• A range of solutes, including polyethylene glycols and salts, were tested.
• The influence of solute molecular weight and solution osmolality was examined.

Main Results:

• Solutes were found to prevent the order-to-disorder transition of the BtuB Ton box upon ligand binding, maintaining a more ordered structure.
• The mechanism of solute action was identified as osmotic, involving preferential exclusion from protein cavities and surfaces.
• A sharp molecular weight cutoff was observed for polyethylene glycols, indicating size-dependent exclusion.
• Solutes also induced a more structured conformation in the Ton box when unfolded by detergents or mutations.

Conclusions:

• Solutes can modulate conformational changes in BtuB and potentially other outer membrane transporters through osmotic effects.
• Hydration changes play a significant role in regulating transporter structure and function.
• These findings have implications for understanding transporter mechanisms and for protein crystallization techniques.