Structural Changes and Quantification of Ligand Binding Affinity in Membrane Transport Proteins

Membrane transport proteins play a very significant role of transporting biosynthesis raw materials, metabolism byproducts and drug molecules across the cell membrane. The Major Facilitator Superfamily (MFS) is an important class of membrane transporters whose members are found in unicellular organism to complex systems like the human central nervous system. Lactose Permease (LacY) of E. coli is an important MFS protein which facilitates transport of various sugar molecules across the plasma membrane. Though the structure of LacY open to cytoplasm has been determined, the structure open to periplasm, which is crucial in understanding the detailed mechanism of transport, is unrevealed. Molecular Dynamics Simulations with an explicit lipid bilayer were carried out with two different conformations of LacY obtained from previous implicit lipid bilayer simulations with the Glu²⁶⁹ protonated. Simulations were carried out in NPAT ensemble with a suggested experimental area of 65.2 Ų per lipid as well as an area of 60 Ų per lipid. The helix-helix distances of LacY were compared with the experimental results obtained from DEER experiments (Smirnova et al., PNAS, 2007) and agree favorably with their suggested periplasmic open state. Moreover to quantify the anomeric binding affinity of sugars to LacY, free energies of binding for different sugar molecules were calculated using alchemical free energy perturbation (FEP) method with restraining potentials applied on sugar molecules for efficient sampling. Free energy calculation results were compared with the experimental results. (Nie et al., JBC, 2006)