Observation of a dewetting transition in the collapse of the melittin tetramer

Marked hydration changes occur during the self-assembly of the melittin protein tetramer in water. Hydrophobicity induces a drying transition in the gap between simple sufficiently large (more than 1nm 2 ) strongly hydrophobic surfaces as they approach each other 1‐6 , resulting in the subsequent collapse of the system, as well as a depletion of water next to single surfaces 7‐10 . Here we investigate whether thehydrophobicinduced collapse of multidomain proteins or the formation of protein oligimers exhibits a similar drying transition. We performed computer simulations to study the collapse of the tetramer of melittin in water, and observed a marked water drying transition inside a nanoscale channel of the tetramer (with a channel size of up to two or three water-molecule diameters). This transition, althoughoccurringonamicroscopiclengthscale,isanalogoustoa first-order phase transition from liquid to vapour. We find that this drying is very sensitive to single mutations of the three isoleucines to less hydrophobic residues and that such mutations in the right locations can switch the channel from being dry to being wet. Thus, quite subtle changes in hydrophobic surface topology can profoundly influence the drying transition. We show that, even in the presence of the polar protein backbone, sufficiently hydrophobic protein surfaces can induce a liquid‐vapour transition providing an enormous driving force towards further collapse.This behaviour was unexpected because of the absence of drying in the collapse of the multidomain protein 2,3-dihydroxybiphenyl dioxygenase (BphC). Proteins exert electrostatic forces on water and are therefore much more complicated than purely hydrophobic systems. We have seen previously that because of these electrostatic forces there was no strong drying transition in the collapse of the BphC enzyme 11 , a twodomain protein. These findings showed that realistic protein‐water forces have a significant function in hydrophobic collapse, and argued against a drying-induced reorganization model of protein folding. Others have examined the role of water during protein folding, finding that water can persist inside the core even after it collapses and can also act as a lubricant for folding 12,13 , but those studies did not address the drying transition that we discuss here. A coarse-grained simulation of the collapse of a model hydrophobic polymer chain showing that the evaporation (drying) of water in the vicinity of the polymer provides the driving force for the collapse, and that the rate-limiting step is the nucleation of a sufficiently large bubble, caused its authors 14 to speculate, “Our findings would seem also pertinent to the mechanism of biological assembly, such as protein folding, but to demonstrate so with simulation will require ananalogoussimulationstudyofaprotein-likechain”.Herewestudy the water drying transition inside the melittin tetramer. Melittin, a 26-residue polypeptide, is a small toxic protein found in honeybee venom, which often self-assembles into a tetramer. Previous simulations of hydration structure around the melittin dimer partly guided our choice of the tetramer for study 15 . The two dimers of the melittin tetramer are separated by a certain distance to create a nanoscale water ‘channel’. The enlarged melittin tetramer is then simulated with molecular dynamics. A strong, cooperative water drying transition is found inside the nanoscale channel whose size is large enough to encompass two or three water-molecule diameters, in agreement with previous speculation 14 .