Estimation of the pore size of the large-conductance mechanosensitive ion channel of Escherichia coli

Abstract

The open channel diameter of Escherichia coli recombinant large-conductance mechanosensitive ion channels (MscL) was estimated using the model of Hille (Hille, B. 1968. Pharmacological modifications of the sodium channels of frog nerve. J. Gen. Physiol. 51:199-219) that relates the pore size to conductance. Based on the MscL conductance of 3.8 nS, and assumed pore lengths, a channel diameter of 34 to 46 A was calculated. To estimate the pore size experimentally, the effect of large organic ions on the conductance of MscL was examined. Poly-L-lysines (PLLs) with a diameter of 37 A or larger significantly reduced channel conductance, whereas spermine (approximately 15 A), PLL19 (approximately 25 A) and 1,1'-bis-(3-(1'-methyl-(4,4'-bipyridinium)-1-yl)-propyl)-4,4'-b ipyridinium (approximately 30 A) had no effect. The smaller organic ions putrescine, cadaverine, spermine, and succinate all permeated the channel. We conclude that the open pore diameter of the MscL is approximately 40 A, indicating that the MscL has one of the largest channel pores yet described. This channel diameter is consistent with the proposed homohexameric model of the MscL.

The gating mechanism of the bacterial mechanosensitive channel MscL revealed by molecular dynamics simulations

“…However, this value (5.8 Å) is much smaller than the open pore size estimated by electrophysiological analyses or channelmediated protein efflux measurements. 6,43,44 This suggests that the result here may reflect an initial conformational change on the way toward the full opening of MscL, which will be discussed later.…”

Hydrophobic gating of mechanosensitive channel of large conductance evidenced by single-subunit resolution

“…The closed channel has a pore diameter of about 3.5 Å (3). The estimated open pore diameter, on the other hand, is 30-40 Å (4,5), suggesting that significant conformational changes take place upon gating.…”

Bacterial Mechanosensitive Channels: Models for Studying Mechanosensory Transduction

“…The pore closure is exposed on the periplasmic side, while on the cytoplasmic side, the pore is shielded by the five C-terminal helices (24,114). The pore region of MscL lacks any charged residues for imparting an ion selectivity filter function and, by opening a pore of *30 Å in diameter, expands to such a degree that ion selection becomes impossible in the open state, rendering the channel nonselective (31,38,61,121). This lack of ion selectivity in MscL and the large size of its open pore are essential for allowing passage of large organic osmolytes of *1000 in molecular weight and, hence, fulfilling its physiological role as a highly efficient emergency valve to release solutes from bacterial cells under .…”

“…MscL produces a nonselective pore that is *3 nm in diameter when activated (33,38), and it has been shown that…”

“…They could then be forced into opening by the application of select methanethiosulfonate reagents (161,165). MscL is well suited for this type of research because of its very large pore diameter and lack of ion selectivity (38), enabling molecules such as the fluorescent Alexa-Fluor molecules (*700 Da) and fluorescently labeled phalloidin (*1.3 kDa) to be delivered into live mammalian cells (41). Being able to transfect mammalian cells with this MscL mutant will open up exciting new possibilities for live-cell fluorescent staining and for the controlled intracellular delivery of various ligands.…”