INVERSION OF RECEPTOR-BINDING PREFERENCES BY MUTAGENESIS - FREE-ENERGY THERMODYNAMIC INTEGRATION STUDIES ON SUGAR BINDING TO L-ARABINOSE BINDING-PROTEINS

The Escherichia coli L-arabinose-binding protein (ABP) participates as a specific receptor in the transport of L-arabinose, D-fucose, and D- galactose through the periplasmic space. The wild-type protein binds L -arabinose about 40 times more strongly than D-fucose. A mutation of t he protein at position 108 (Met --> Leu) causes a specificity change. The Met108Leu ABP slightly prefers binding of D-fucose over L-arabinos e. Molecular dynamics (MD) and thermodynamic integration (TI) computer simulations were performed to study the mechanism of sugar discrimina tion and specificity change based on the known high-resolution X-ray s tructures. The specificity change was evaluated by calculating the dif ference in free energy of L-arabinose versus D-fucose bound to wild-ty pe and Met108Leu ABP. The calculated free energy differences are consi stent with the experimentally observed specificity of wild-type and Me t108Leu ABP. The simulations indicate that the specificity change of M et108Leu ABP is accomplished mainly by reduced Lennard-Jones interacti ons of residue 108 with L-arabinose and improved interactions with D-f ucose. In addition to MD/TI calculations on sugar binding, finite diff erence Poisson-Boltzmann calculations were performed to identify the m ost stable ionization state of buried ionizable residues in ABP.