The Role of MscS Cytoplasmic Domain As An Osmolyte Filter

Abstract

Mechanosensitive (MS) channels, inner membrane proteins of bacteria, open and close in response to mechanical stimuli such as changes in membrane tension during osmotic stress. In bacteria, these channels act as safety valves thus preventing cell rupture upon hypoosmotic shock.The MS channels of small conductance, MscS, are homoheptameric and consist of a large cytoplasmic (CP) domain that features a balloon-like, water filled chamber opening to the cytoplasm through seven side pores and a small distal pore. This CP domain is considered to be a molecular sieve, which prevents a loss of essential osmolytes and metabolites at the cytoplasmic side. In bacteria, glutamate is a predominant anion that helps to maintain potassium pools at an optimum level and also is a prevalent osmoprotectant maintaining the cell turgor. We have explored, using molecular simulations, the free energy landscape characterizing the translocation and exit of a glutamate ion through one of the side pores, to illustrate the role of the CP domain in selective filtering of glutamate. The adaptive biasing force (ABF) method is applied to the glutamate molecule to determine the free energy barrier along the chosen translocation pathway. The transport kinetics of glutamate based on the measured free energy profile, suggests the presence of an entropic barrier that slows down the passage of glutamate through the pore but lets water pass quickly. A low enthalpic barrier of ∼4 kBT is sufficiently low to prevent glutamate from clogging the pore. Analysis of the electrostatic potential of the CP domain indicates that the CP interior presents an environment relatively unfavorable for anions.