1. H v 1: How A Voltage-sensor May Form A Channel Younes Mokrab Biophysical Society 53 rd Annual Meeting 2 Mar 2009

2. H v 1: a VS with proton channel activity Ramsey et al. 2006 Schilling et al. 2002 H+H+ S1S2 S3S4 + + + Exact function not known but enriched in immune tissue. Topology similar to VS domains of voltage-gated ion channels. In response to depolarising potential, it conducts an outward H + current, with similar characteristics to G vH+. Conduction pore and molecular mechanism is still unknown I step -I tail

3. Homology open-state models for H v 1 and simulation of insertion into bilayer Bond et al. 2006, Jiang et al. 2002, Jiang et al. 2003, Long et al. 2007 4:1 Full-atom Coarse-grain Protein Lipid Water Ion H v 1 homology models based on x-ray structures of K v AP, K v 1.2 and K v 1.2-2.1 chimera. Self assembly in POPC for 320 ns Protein inserted to membrane Coarse-grain (CG) models

4. Full-atom simulations based on CG configuration of protein in bilayer Per-residue fluctuationBackbone RMSD versus time Conversion of system to full-atom using LSF Full-atom simulation for 20 ns

5. Hydrophilic residues cluster along the centre of the H v 1 bundle Chimera-based K v 1.2-basedk v AP-based Models might capture different open-state conformations, all showing charged/polar residues which line a potential conduction pathway for H + during H v 1 activation.

6. A water-permeable pore forms along central axis of H v 1 Analysis of water density during simulation shows water penetration of Hv1 along its central axis in the models, forming a potential water permeable pore. Chimera-based K v 1.2-basedk v AP-based

7. Water molecules are coordinated by polar and charged residues A number of charged side chains are identified which may be involved in direct H + translocation, and/or polar side chains in general might coordinate a ‘proton wire’. In silico and In vivo mutagenesis experiment in progress to test the roles of specific residues.

8. Water pore absent from VS homologues Chimera K v 1.2k v AP Simulations of other voltage-gated channels show considerably-reduced water penetration along the central axis of the VS domains.

9. Pore-lining residues unique to H v 1

10. Conclusions and future directions Combined CG-AT simulations of H v 1 open-state models in solvated lipid bilayer reveals a central water-permeable pore which seems to be absent from other VS proteins. High water density sites in a potential ‘gating pore’ in H v 1 appear to be coordinated by uniquely conserved polar and charged residues. Role of specific residues in water coordination is being tested by in silico and in vivo site-directed mutagenesis.

11. Acknowledgments Professor Mark Sansom, SBCB, University of Oxford. Dr Scott Ramsey and Professor David Clapham, Harvard Medical School (Poster 3415-Pos B462, Wed 4 Mar 1-3 pm). Dr Zara Sands, AstraZeneca, Sweden. Dr Kathryn Scott, Dr Phil Stansfeld, Dr Kaihsu Tai and members of SBCB.