Probing the stability and interdomain interactions in the ABC transporter OpuA, using single-molecule optical tweezers

Abstract

Transmembrane transporter proteins are essential for the maintenance of cellular homeostasis and, as such, are key drug targets. Many transmembrane transporter proteins are known to undergo large structural rearrangements during their functional cycles. Despite the large amount of detailed structural and functional data available for these systems, our understanding of their dynamics and therefore how they function is generally limited. We introduce an innovative approach which enables us to directly measure the dynamics and stabilities of inter-domain interactions of transmembrane proteins using optical tweezers. Focusing on the osmoregulatory ABC transporter OpuA from Lactococcus lactis, we examine the mechanical properties and potential interactions of its substrate-binding domains. Our measurements are performed in lipid nanodiscs, providing a native-mimicking environment for the transmembrane protein. The technique provides high spatial and temporal resolution and allows us to study the functionally-relevant motions and inter-domain interactions of individual transmembrane transporter proteins in real-time in a lipid bilayer.