The role of MscL amphipathic N terminus indicates a blueprint for bilayer-mediated gating of mechanosensitive channels

Abstract

The bacterial mechanosensitive channel MscL gates in response to membrane tension as a result of mechanical force transmitted directly to the channel from the lipid bilayer. MscL represents an excellent model system to study the basic biophysical principles of mechanosensory transduction. However, understanding of the essential structural components that transduce bilayer tension into channel gating remains incomplete. Here using multiple experimental and computational approaches, we demonstrate that the amphipathic N-terminal helix of MscL acts as a crucial structural element during tension-induced gating, both stabilizing the closed state and coupling the channel to the membrane. We propose that this may also represent a common principle in the gating cycle of unrelated mechanosensitive ion channels, allowing the coupling of channel conformation to membrane dynamics.

Spectator no more, the role of the membrane in regulating ion channel function

“…FRET [20] measurements of E. coli MscL opened by addition of lyso-PC estimated a pore with over 25Å in diameter and favoured a helix-tilt gating model [20]. A combination of cwEPR, electrophysiology measurements and computational studies highlighted the importance of the N-terminal amphipathic S1 helix on MscL gating [21] (Fig 2a), a region identified to interact with lipids [22]. Most recently two crystal stuctures of an archeal MscL orthlogue [23] revealed two states, one closed (pore diameter around 4Å) and the other at least partially opened (pore diameter around 8Å).…”

Structural Dynamics of the MSCL C-Terminal Domain

“…We have reported previously that distortion of transbilayer pressure profiles through the use of mixtures of lipids with different geometries leads to open MscL channels in patch-clamp experiments with no suction applied to the patch pipette 6,8,20,35 . A comparison of EPR results obtained from closed versus open channels (in the absence and the presence of 25% molar LPC respectively) is shown in Fig.…”

“…Moreover, the open channel structure was also determined by SDSL EPR spectroscopy 8 allowing for estimation of the size of the open channel pore in a very good agreement with electrophysiological sieving studies 9 . Later on, an improved open E. coli MscL (EcMscL) 3D structure was determined using both ensemble and single molecule site-directed fluorofore labelling (SDFL) FRET spectroscopy in combination with MD simulations 6,10,11 in agreement with the X-ray structure of an expanded form of an archaeal MscL homolog from Methanosarcina acetivorans 12 . While the overall gating-related structural changes in MscL have largely been established 8,11,13,14 the structural dynamics and physiological role of the C-terminal domain has thus far been controversial.…”

“…These points do not have any properties other than position. The lipid bilayer was assumed to behave as a linear elastic plate 27,28 , and the secondary structural elements making the MscL protein were treated as elastic cylinders/rods, as described 6,29,30 . The Young's modulus of the bilayer was assumed to be 4.3 MPa as previously determined using excised patch fluorometry 28 and the Young's moduli of the MscL N-terminal (0.35 GPa), TM1 (2.6 GPa), TM2 (3.4 GPa) and C-terminal (7.7 GPa) α-helices were determined using steered molecular dynamics (MD) simulations 31 .…”

“…The three-dimensional (3D) structure of MscL from Mycobacterium tuberculosis (MtMscL) in its closed conformation was resolved at 3.5 Å 3 . The channel is a homopentamer consisting of N-and C-terminal domains facing the cytoplasm and TM1 and TM2 transmembrane helices connected by a periplasmic loop [3][4][5][6] . The closed channel structure was also determined in the lipid bilayer by site-directed spin labelling electron paramagnetic resonance (SDSL EPR) spectroscopy 7 .…”