Conformational Transitions Of Uracil Transporter UraA From Escherichia Coli:A Molecular Simulation Study

Coupled tansporter mediates all kinds of ions and molecules across a cell membrane.NSC2 family proteins play an important role in the physiological activities of the various kinds of organisms.Representative NAT/NCS2 family proteins include the vitamin C transporters SVCT1 and SVCT2 in human being;the uracil transporter rSNBT1 in mammals and the Escherichia coli uracil/H~+symporter UraA.Transporters of the same family protein are homologous in sequence and structure.Therefore,the in-depth study of homologous protein UraA can also provide a theoretical basis for future related NAT family proteins in humans.Along with the development of the computer technology and the calculation methods,molecular dynamics method begins to be applied to the exploration of biological macromolecular structure system at the molecular and atomic level.It can explain activity regularity of protein structure and function mechanism.Some experiments proposed a working model to explain proton-coupling and uracil transporting process of UraA on the basis of the crystal structure of UraA,but the details of conformational changes remained unknown.Thus,in order to make clear the critical factors in conformational changes from inward-open to outward-open.Five MD simulations were used to study the conformation of UraA complexes in different protonation states.From our known results,UraA must be able to adopt an outward-open conformation,it is convenient and effective to combine the extracellular substrate uracil and proton.Then this is probably accomplished by a rigid-body rotation of the gata domain to the core domain.The conformation changes from outward-open to inward-open.And the conformation would come back to outward-open with the transfer of proton and the release of the substrate.It gives rise to significant structural rearrangements of H-bond network.Those H-bonds contribute to stabilization of UraA.We analysed essential amino acids in H-bond network.It can be concluded that Glu290 was crucial in a network of hydrogen-bonds in the middle of the core domain involving another essential residue,mainly including Glu241,His245,Tyr288,Tyr342,Ser338,and the network of hydrogen-bonds was the key to maintain the stability of conformation.Protonation of Glu290 affects the stability of network of H-bond and changed the transmembrane segments of TM3,TM10 and TM12.Thus,Glu290 may play a vital role as“proton trigger”that affects spatial structural of amino and residues near substrate binding side leading to an outward-open conformation transition.