Small Substrate Transport and Mechanism of a Molybdate ABC Transporter in a Lipid Environment [Membrane Biology]

April 10th, 2014 by Rice, A. J., Harrison, A., Alvarez, F. J. D., Davidson, A. L., Pinkett, H. W.

Embedded in the plasma membrane of all bacteria, ABC importers facilitate the uptake of several vital nutrients and cofactors. The ABC transporter, MolBC-A imports molybdate by passing substrate from the binding protein MolA to a membrane-spanning translocation pathway of MolB. To understand the mechanism of transport in the biological membrane as a whole, the effects of the lipid bilayer on transport needed to be addressed. CW-EPR and in vivo molybdate uptake studies were used to test the impact of the lipid environment on the mechanism and function of MolBC-A. Working with the bacterium Haemophilus influenzae, we found that MolBC-A functions as a low affinity molybdate transporter in its native environment. In periods of high extracellular molybdate concentration, H. influenzae makes use of parallel molybdate transport systems (MolBC-A and ModBC-A) to take up a greater amount of molybdate than a strain with ModBC-A alone. In addition, the movement of the translocation pathway in response to nucleotide binding and hydrolysis in a lipid environment is conserved when compared to in-detergent analysis. However, EPR spectroscopy indicates that a lipid environment restricts the flexibility of the MolBC translocation pathway. By combining CW-EPR spectroscopy and substrate uptake studies, we reveal details of molybdate transport and the logistics of uptake systems that employ multiple transporters for the same substrate, offering insight into the mechanisms of nutrient uptake in bacteria.