In hydrophilic protein-protein associations, the dehydration penalty, which can cause the formation of a reaction barrier, must be canceled out; however, its mechanism has not been clarified. Here, we explored the possible mechanism through investigation of the dimerization of nucleotide binding domains (NBDs). We assessed the different dimerization processes by molecular dynamics simulations with and without thermal fluctuations in each NBD. Consequently, the reaction barriers of the former and latter were estimated to be similar to 100 and similar to 15 kcal/mol, respectively, suggesting that thermal fluctuations in the proteins facilitate the exclusion of water molecules from the interfacial region, thereby lowering the barrier. (C) 2015 Elsevier B.V. All rights reserved.