Abstract
The kinetic Monte Carlo reaction-path-following technique is applied to determine the lowest-energy water pathway and the coordinating amino acids in bAQP1 and GlpF channels, both treated as rigid. In bAQP1, water molecules pass through the pore between the asparagine-proline-alanine (NPA) and selectivity filter (SF) sites one at a time. The water chain is interrupted at the SF where one water forms three stable hydrogen bonds with protein atoms. In this SF, water's conformation depends on the protonation locus of H182. In GlpF, two water molecules bond simultaneously to the NPA asparagines and pass through the SF in zigzag fashion. No water single-file forms in rigid GlpF. To accommodate a single file of waters requires narrowing the GlpF pore. Our results reveal that in both proteins a proposed bipolar water arrangement is thermally disrupted in the NPA region, especially in the cytoplasmic part of the pore. The equilibrium hydrogen-bonded chain is occasionally interrupted in the hydrophobic zones adjacent to the NPA motifs. The permeation of alkali cations through bAQP1 and GlpF is barred due to a large free-energy barrier in the NPA region as well as a large energy barrier blocking entry from the cytoplasm. Permeation of halides is prevented due to two large energy barriers in the cytoplasmic and periplasmic pores as well as a large free-energy barrier barring entry from the periplasm. Our results, based on modeling charge permeation, support an electrostatic rather than orientational basis for proton exclusion. Binding within the aquaporin pore cannot compensate sufficiently for dehydration of the protonic charge; there is also an electrostatic barrier in the NPA region blocking proton transport. The highly ordered single file of waters, which is drastically interrupted at the SF of bAQP1, may also contribute to proton block.