An important focus of our laboratory is the understanding of the structure and function of ion channels. We are particularly interested in issues about ion permeation, ion selectivity, gating, and channel inhibitors. We are currently working on the KcsA channel, the OmpF porin, and the gramicidin A> channel. We are also spending our efforts in the development of new computational approaches for studying biological macromolecular systems.

The computational approach called "molecular dynamics" (MD) is central to our work. It consists of constructing detailed atomic models of the macromolecular system and, having described the microscopic forces with a potential function, using Newton's classical equation, F=MA, to literally "simulate" the dynamical motions of all the atoms as a function of time. The calculated trajectory, though an approximation to the real world, provides detailed information about the time course of the atomic motions, which is impossible to access experimentally.

In addition, other computational approaches, at different level of complexity and sophistication, can be very useful. In particular, Poisson-Boltzmann (PB) continuum electrostatic models, in which the influence of the solvent is incorporated implicitly, plays an increasingly important role in estimating the solvation free energy of macromolecular assemblies.

Research topics:

• KcsA
  The determination of the structure of the KcsA K+ channel from Streptomyces lividan has made it possible to investigate the function of a biological channel at the atomic level ( Doyle et al, 1998 ). Because of its structural similarity with eukaryotic K-channels, investigations of KcsA are expected to help understand a large class of biologically important channels. We are currently working on several fundamental aspect of the function of the KcsA channel such as, electrostatics, muti-ion permeation, channel gating, and inhibition by various blocking agents (see Roux and MacKinnon, 1999 ; Berneche and Roux, 2000 ; Roux et al., 2000 ; Crouzy, et al, 2001; ; Berneche and Roux, 2001 ).



• OmpF porin
  The outer membrane of Escherichia coli protects the cell against hostile agents and facilitates the uptake of nutrients. This activity is mediated by macromolecular structures called porins. Porins are not very selective and have only some specificity towards cations or anions. The porins represent ideal systems for addressing the fundamental electrostatics principles governing ion flow in molecular pores. Because high resolution well-characterized structures are available for outer membrane porins of E coli, they provide excellent model channel systems for computations based on detailed atomic models. We are currently working on the cation-selective matrixporin (OmpF) which is a major component of the E. coli outer membrane.



• Brownian dynamics
  Brownian dynamics provides an attractive computational approach for simulating the permeation of ions over long time-scales without having to treat all the solvent molecules explicitly. The approach consists in integrating stochastic equation of motions with some effective potential which incorporates the systematic influence of the environment. To account for non-equilibrium boundary conditions found in ion channel systems (asymetric ion concentration and transmembrane potential) we have developed a method combining the Grand Canonical Monte Carlo algorithm with Brownian dynamics, GCMC/BD ( Im et al., 2000 ).



• Implicit solvation methods
  MD simulations with explicit solvent molecules are computationally expensive and important properties such as solvation free energies may often converge slowly. Other computational approaches in which the influence of the solvent is incorporated implicitly are needed (see Roux and Simonson, 1999 ). Those include continuum electrostatics based on the Poisson-Boltzmann (PB) equations ( Nina et al., 1997 ), stochastic Brownian Dynamics ( Im et al., 2000 ), , and mean-field theories based on statistical mechanical integral equations ( Beglov and Roux, 1997 ; Du et al., 2000 ).



• Polarizable force field
  The potential function is one of the most important ingredients in MD calculations. Many of the current simulations of biomolecular systems are based on a potential function representing the interactions between non-bonded atoms in terms of a Lennard-Jones 6-12 potential and fixed atomic partial charge coulomb electrostatics. In our group we normally use the potential function of the CHARMM biomolecular simulation program. Other similar potential functions are AMBER , and GROMOS . We think that such simple models, which ignore electronic polarization effects, will be insufficient for understanding the microscopic basis of ion selectivity in biological channels. We are currently developing a force field which will include induced polarization.



• Free energy simulations
  By and large, microscopic processes such as ion permeation, macromolecular conformational changes, ligand binding specificity, and protein-protein association are driven thermodynamically by the free energy (or potential of mean force) in diverse and complex environments such as bulk aqueous solution, the active site of an enzyme, the interior of an ion channel, or a bilayer membrane. A quantitative determination of free energies is thus a problem of central importance in theoretical biophysics. We are currently developing and extending current methodologies to allow precise and computationally inexpensive free energy calculations (see Pomes et al., 1999 , Shobana et al., 2000 ).