A Light-Actuated Nanovalve Derived from a Channel Protein

Koçer, Armağan; Walko, Martin; Meijberg, Wim; Feringa, Ben L.

doi:10.1126/science.1114760

Cited by 509 publications

(463 citation statements)

References 25 publications

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“…2) (16). Even though in nature, MscL opens in response to tension in the lipid bilayer, attaching charged cysteine-specific compounds to the pore of MscL-G22C homopentamers opens the channel spontaneously (36)(37)(38). Recently, based on this principle, we gained control on the degree of MscL opening in the absence of tension.…”

Section: Ion Mobility-mass Spectrometry Can Distinguish Individual Hementioning

confidence: 99%

Global structural changes of an ion channel during its gating are followed by ion mobility mass spectrometry

Konijnenberg

Yilmaz²,

Ingólfsson

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Mechanosensitive ion channels are sensors probing membrane tension in all species; despite their importance and vital role in many cell functions, their gating mechanism remains to be elucidated. Here, we determined the conditions for releasing intact mechanosensitive channel of large conductance (MscL) proteins from their detergents in the gas phase using native ion mobilitymass spectrometry (IM-MS). By using IM-MS, we could detect the native mass of MscL from Escherichia coli, determine various global structural changes during its gating by measuring the rotationally averaged collision cross-sections, and show that it can function in the absence of a lipid bilayer. We could detect global conformational changes during MscL gating as small as 3%. Our findings will allow studying native structure of many other membrane proteins.ion mobility mass spectrometry | MscL | membrane proteins | structure function | ion channel gating O ne of the best candidates to explore the gating of mechanosensitive channels is the mechanosensitive channel of large conductance (MscL) from Escherichia coli. The crystal structure of MscL in its closed/nearly closed state from Mycobacterium tuberculosis revealed this channel as a homopentamer (1). Each subunit has a cytoplasmic N-and C-terminal domain as well as two α-helical transmembrane (TM) domains, TM1 and TM2, which are connected by a periplasmic loop. The five TM1 helices form the pore and the more peripheral TM2 helices interact with the lipid bilayer.MscL detects changes in membrane tension invoked by a hypoosmotic shock and couples the tension sensing directly to large conformational changes (1, 2). On the basis of a large body of structural and theoretical data, numerous gating models of MscL have been proposed (3-9). These models agree upon (i) the hydrophobic pore constriction of the channel and (ii) the channel opens by an iris-like rotation-i.e., a tilting and outward movement of transmembrane helices that make the channel wider and shorter (5). This mechanism is supported by patchclamp (10), disulfide cross-linking (11), FRET spectroscopy (12), and site-directed spin labeling EPR experiments (6, 7), as well as computational studies (13-15). So far, direct experimental results have only been observed for short-range local structural changes, and no measure of the overall global structural changes during channel gating have been reported. Because there is no crystal structure available for the open MscL channel, elucidating overall global structural changes from the onset of channel activation is of utmost importance for our understanding of the gating mechanism of mechanosensitive channels. Here, we provide direct experimental evidence for the key areal changes occurring during channel gating by combining our ability to activate MscL in a controlled manner to different subopen states (16) with a native ion mobility-mass spectrometry (IM-MS) approach. Results and DiscussionDetergents that are Unstable in the Gas Phase Allow Detection of MscL in Its Native Form. Native mass ...

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Section: Ion Mobility-mass Spectrometry Can Distinguish Individual Hementioning

confidence: 99%

Global structural changes of an ion channel during its gating are followed by ion mobility mass spectrometry

Konijnenberg

Yilmaz²,

Ingólfsson

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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“…Light-driven changes in molecular polarity can be used to power the opening and closing of a nanovalve (71). Figure 11 illustrates the use of the spiropyran-mercyanine system for this purpose.…”

Section: Photoswitchable Systems In Lipid Bilayersmentioning

confidence: 99%

Light Switching of Molecules on Surfaces

Browne

Feringa

2009

Annu. Rev. Phys. Chem.

Self Cite

274

254

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Smart surfaces, surfaces that respond to an external stimulus in a defined manner, hold considerable potential as components in molecular-based devices, not least as discrete switching elements. Many stimuli can be used to switch surfaces between different states, including redox, light, pH, and ion triggers. The present review focuses on molecular switching through the electronic excitation of molecules on surfaces with light. In developing light-responsive surfaces, investigators face several challenges, not only in achieving high photostationary states and fully reversible switching, but also in dealing with fatigue resistance and the effect of immobilization itself on molecular properties. The immobilization of light-responsive molecules requires the design and synthesis of functional molecular components both to achieve light switching and to anchor the molecular entity onto a surface. This review discusses several demonstrative examples of photoswitchable molecular systems in which the photochemistry has been explored in the immobilized state under ambient conditions and especially on electroactive surfaces, including self-assembled monolayers, bilayers, and polymer films.

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“…An alternative technique, allowing the study of bulk fluxes mediated by MscL in reconstituted lipid vesicles, was introduced by Kocer et al (20,21). This involved trapping high concentrations of the fluorescent dye calcein within the lumen of the vesicles, so that the intensity of calcein fluorescence was reduced by concentration quenching.…”

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confidence: 99%

Importance of Direct Interactions with Lipids for the Function of the Mechanosensitive Channel MscL

2008

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We have studied the effects of lipid structure on the function of the mechanosensitive channel of large conductance (MscL) from Escherichia coli to determine whether effects follow from direct interaction between the lipids and protein or whether they follow indirectly from changes in the curvature stress in the membrane. The G22C mutant of MscL was reconstituted into sealed vesicles containing the fluorescent molecule calcein, and the release of calcein from the vesicles was measured following opening of the channel by reaction with [2-(triethylammonium)ethyl] methanethiosulfonate (MTSET), which introduces five positive charges into the region of the pore constriction. The presence of anionic lipids in the vesicle membrane changed the rates and amplitudes of calcein release, the effects not correlating with calculated changes in lipid spontaneous curvature. Mutation of charged residues in the Arg-104, Lys-105, Lys-106 cluster removed high-affinity binding of anionic lipids to MscL, and the presence of anionic lipid no longer affected calcein flux through MscL. Changing the zwitterionic lipid from phosphatidylcholine to phosphatidylethanolamine resulted in a large decrease in the rate of calcein release, the change in rate varying linearly with lipid composition, as expected if spontaneous curvature affected the rate of release. However, rates of release of calcein measured in the presence of phosphatidylethanolamine-N-methyl and phosphatidylethanolamine-N,N-dimethyl did not fit the correlation between rate and curvature established for the phosphatidylcholine/phosphatidylethanolamine mixtures. Rather, the effects of zwitterionic lipid headgroup on calcein flux suggested that what was important was the presence of a proton in the headgroup, able to take part in hydrogen bonding to MscL. We conclude that the function of MscL is likely to be modulated by direct interaction with the surrounding, annular phospholipids that contact the protein in the membrane.

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A Light-Actuated Nanovalve Derived from a Channel Protein

Cited by 509 publications

References 25 publications

Global structural changes of an ion channel during its gating are followed by ion mobility mass spectrometry

Global structural changes of an ion channel during its gating are followed by ion mobility mass spectrometry

Light Switching of Molecules on Surfaces

Importance of Direct Interactions with Lipids for the Function of the Mechanosensitive Channel MscL

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