Abstract
All-atom NMA is used to explore possible mechanisms for slow gating in ClC Cl− channels. As the “double-barreled” architecture is well established throughout the ClC family, both channels and transporters (Miller, 2006. Nature. 440:484), we use the high-resolution (2.5 Å) X-ray structure of an E. coli ClC transporter (pdb entry 1OTS) as a template, describe it with the CHARMM22 force field and carry out standard all-mode NMA. The slowest, intrinsic motions encoded in the structure are determined by protein shape. Perturbing the system in either direction along the 7th all-atom NM leads to slow relative swinging of the subunits, perpendicular to the membrane plane. The in-plane swivel axis lies at the subunit interface, near the protein's center. The intracellular interfacial domain is the region most affected. Here the two halves of the protein oscillate, separating and then nearly touching. The R and A helices execute large scale swaying, alternately increasing and decreasing their cytoplasmic ends’ separations, motion in agreement with FRET experiments (Bykova et al., 2006. Nat. Struct. Biol. 13:1115). The ion-occupied intracellular pores behave as almost rigid units. As the subunits separate, the intracellular pore tilt relative to the membrane plane changes notably. In contrast, the extracellular portion of the subunit interface is significantly less affected, although small interfacial structural changes are clearly observable. Those extracellular regions structurally affected by the subunits’ slow sway are localized near the extracellular Cl− pathways. As the subunits separate, these regions compress, possibly shutting the extracellular pores. As they approach, the extracellular regions near the Cl− conduction pathways relax, possibly opening them.