title: Molecular dynamics investigation of Cl- and water transport through a eukaryotic CLC transporter.
abstract: Early crystal structures of prokaryotic CLC proteins identified three Cl(-) binding sites: internal (S(int)), central (S(cen)), and external (S(ext)). A conserved external GLU (GLU(ex)) residue acts as a gate competing for S(ext). Recently, the first crystal structure of a eukaryotic transporter, CmCLC, revealed that in this transporter GLU(ex) competes instead for S(cen). Here, we use molecular dynamics simulations to investigate Cl(-) transport through CmCLC. The gating and Cl(-)/H(+) transport cycle are inferred through comparative molecular dynamics simulations with protonated and deprotonated GLU(ex) in the presence/absence of external potentials. Adaptive biasing force calculations are employed to estimate the potential of mean force profiles associated with transport of a Cl(-) ion from S(ext) to S(int), depending on the Cl(-) occupancy of other sites. Our simulations demonstrate that protonation of GLU(ex) is essential for Cl(-) transport from S(ext) to S(cen). The S(cen) site may be occupied by two Cl(-) ions simultaneously due to a high energy barrier (∼8 Kcal/mol) for a single Cl(-) ion to translocate from S(cen) to S(int). Binding two Cl(-) ions to S(cen) induces a continuous water wire from S(cen) to the extracellular solution through the side chain of the GLU(ex) gate. This may initiate deprotonation of GLU(ex), which then drives the two Cl(-) ions out of S(cen) toward the intracellular side via two putative Cl(-) transport paths. Finally, a conformational cycle is proposed that would account for the exchange stoichiometry.
authors.affiliation: Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
authors.forename: Mary Hongying
authors.initials: MH
authors.lastname: Cheng
authors.forename: Rob D
authors.initials: RD
authors.lastname: Coalson
mesh_terms: D001665:Binding Sites; D001692:Biological Transport; D018118:Chloride Channels / Q000737:chemistry / Q000378:metabolism; D002712:Chlorides / Q000378:metabolism*; D005057:Eukaryotic Cells / Q000378:metabolism*; D018698:Glutamic Acid / Q000378:metabolism; D042541:Intracellular Space / Q000378:metabolism; D015640:Ion Channel Gating; D007477:Ions; D056004:Molecular Dynamics Simulation; D055550:Protein Stability; D017433:Protein Structure, Secondary; D011522:Protons; D000461:Rhodophyta / Q000378:metabolism*; D012996:Solutions; D013816:Thermodynamics; D013997:Time Factors; D014867:Water / Q000378:metabolism*
chemical_list: D018118:Chloride Channels; D002712:Chlorides; D007477:Ions; D011522:Protons; D012996:Solutions; D014867:Water; D018698:Glutamic Acid