Pore Mutations of the Escherichia coli MscS Channel Affect Desensitization but Not Ionic Preference

Abstract

Mechanosensitive channels rescue bacterial cells from a fate of lysis when they transfer from a high- to low-osmolarity environment. Of three Escherichia coli mechanosensitive proteins studied to date, only MscS-Ec demonstrates a small anionic preference and a desensitized, nonconducting state under sustained pressure. Little is known about the mechanisms generating these distinctive properties. Eliminating the sole positive charge in the MscS-Ec pore region (Arg88) did not alter anionic preference. Adding positive charges at either end of the pore did not augment anionic preference, and placing negative charges within the pore did not diminish it. Thus, pore charges do not control this characteristic. However, from this analysis we identified mutations in the hinge region of the MscS-Ec pore helix (at Gly113) that profoundly affected ability of the channel to desensitize. Substitution with nonpolar (Ala, Pro) or polar (Asp, Arg, Ser) residues inhibited transition to the desensitized state. Interestingly, Gly113 replaced with Met did not impede desensitization. Thus, although Gly is not specifically required at position 113, MscS desensitization is strongly influenced by the residue situated here. Mutations at residues further into the pore also regulated desensitization. Transition to this unique mechanosensitive channel state is discussed in terms of existing data.

Structure and molecular mechanism of an anion-selective mechanosensitive channel of small conductance

“…5B). Previous studies indicate that the β-barrel in EcMscS is important for channel activity (27,31,32). Comparison of the closed and open structures of EcMscS shows that there is no striking difference in the radius of the β-barrel pore size (∼2 Å in radius) (Fig.…”

Selectivity mechanism of the mechanosensitive channel MscS revealed by probing channel subconducting states

“…The cytoplasmic domain is also considered to have a role in the adaptation and inactivation of the channel 25,26,29 and has been suggested to serve as a sensor for cytoplasmic crowding 19,30 . Furthermore, it has been proposed that the MscS cytoplasmic vestibulum may have the role of an ion-selectivity filter in this channel 6,20,31,32 .…”

Bacterial Mechanosensitive Channels: Models for Studying Mechanosensory Transduction

“…However, MscS requires less tension for opening compared with MscL (65,110,144), allowing bacterial cells to respond in a graded manner to hypo-osmotic challenges (16). Unlike MscL, which has no preference for any ions, MscS exhibits a weak preference for anions compared with cations (43,95,140). This weak ion selectivity has recently been shown to originate from the charged residues within seven vestibular portals in the MscS cytoplasmic chamber, which is interesting, because, unlike voltage-gated K + , Na + and Ca 2 + channels, the selectivity of MscS is not determined by charged residues in the channel pore but by residues outside the pore in the waterfilled cytoplasmic domain (36,47,170).…”