State-Dependent and Mutation-Induced Differences in Protein-Lipid Interactions in the Na,K Atpase

Abstract

Specific protein-lipid interactions have emerged as allosteric regulators of membrane protein function. However, the intrinsic dynamic nature of both the protein and the surrounding membrane render structural and biophysical characterization of such interactions are extremely challenging. The sodium-potassium transporter (Na,K ATPase) helps maintain critical electrochemical gradients by an ATP-dependent switch of three intracellular Na+ ions and two extracellular K+ ions across the membrane. Recent experimental findings have pinpointed a few locations of protein-lipid interactions of functional importance. However, a detailed description of how lipid interactions change during the Na,K ATPase membrane switch, in particular in disease mutant forms of the proteins, is currently lacking. Here, we used all-atom molecular dynamics simulations to characterize interactions by annular lipids to sodium- and potassium-bound states of Na,K ATPase in an asymmetric, multicomponent mimic of the plasma membrane. We specifically identified lipid dynamics at a site in the groove between transmembrane domain 3,5 and 7 that has been suggested to regulate function by inhibition and which is also a hot-spot for disease mutations. This presumed inhibitory site showed state-dependent protein-lipid interactions; while POPE lipids and cholesterol embedded the sodium-bound state, the potassium-bound state had a higher presence of POPS and POPI lipids. Upon exploring disease mutant simulations at the protein-lipid interface, we found altered lipid interactions at the mutation sites, in particular we observed stable interactions with PS and PE lipids. These mutant-induced lipid interactions resulted in decreased protein dynamics, which possibly reflect resistance to normal transport behavior. Hence, our simulation approach constitutes a potential tool to better understand shifts in protein-lipid interactions during normal cycling of the Na,K ATPase transporter, and also to better understand the molecular basis of disease mutations.