Kinetics of Gramicidin Channel Formation in Lipid Bilayers: Transmembrane Monomer Association

O'Connell, A. M.; Koeppe, Roger E.; Andersen, Olaf S.

doi:10.1126/science.1700867

Cited by 247 publications

(288 citation statements)

References 30 publications

Supporting

Mentioning

273

Contrasting

Unclassified

Order By: Relevance

“…Moreover, it is well understood why the membrane active form is single-stranded. Indoles are much more stable in the hydrophilic͞hydrophobic bilayer interface than in the hydrophobic core of the bilayer (21,(31)(32)(33). When the tryptophans are completely replaced by phenylalanines, the predominant conformation in the bilayer is the left-handed, double-stranded structure, the same fold as shown in Fig.…”

mentioning

confidence: 52%

Validation of the single-stranded channel conformation of gramicidin A by solid-state NMR

Kovacs

Quine

Cross

1999

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

View full text Add to dashboard Cite

The monovalent cation selective channel formed by a dimer of the polypeptide gramicidin A has a single-stranded, right-handed helical motif with 6.5 residues per turn forming a 4-Å diameter pore. The structure has been refined to high resolution against 120 orientational constraints obtained from samples in a liquid-crystalline phase lipid bilayer. These structural constraints from solid-state NMR ref lect the orientation of spin interaction tensors with respect to a unique molecular axis. Because these tensors are fixed in the molecular frame and because the samples are uniformly aligned with respect to the magnetic field of the NMR spectrometer, each constraint restricts the orientation of internuclear vectors with respect to the laboratory frame of reference. The structural motif of this channel has been validated, and the high-resolution structure has led to precise models for cation binding, cation selectivity, and cation conductance efficiency. The structure is consistent with the electrophysiological data and numerous biophysical studies. Orientational constraints derived from solid-state NMR can be used to determine high-resolution three-dimensional structures. Such an approach has been used to define the structure of the ion channel, gramicidin A, in lamellar phase lipids (ref. 1; PDB accession no. 1MAG). Although a reasonable model of this structure has been extant for nearly 30 years (2) and a structure was determined by solution NMR spectroscopy in SDS micelles (3, 4), crystallographic and solution NMR methods have not been successful in a lipid environment. Recently, the validity of the solid-state NMR structure has been challenged (5). In this report, the structural fold of the channel is validated by comparing predicted and observed values for structural constraints not used quantitatively in solving for the structural fold. Furthermore, the NMR observables are compared with predicted values from several structures in the Protein Data Bank. The results establish the high resolution of the solid-state structure and the clear validity of this motif in a lipid environment.Gramicidin A is a polymorphic structure and the dominant sequence in gramicidin D, the biosynthetic product from Bacillus brevis: HCO-Val-Gly-Ala-DLeu-Ala-DVal-Val-ValTrp-DLeu-Trp-DLeu-Trp-DLeu-Trp-NHCH 2 CH 2 OH. In isotropic organic solvents, this peptide typically forms a doublestranded dimer that may be parallel or antiparallel, lefthanded or right-handed and has a range of residues per turn from 5.6 to 7.2 (5-10). In the heterogeneous anisotropic lipid environment, the structure is almost exclusively singlestranded. Because of the short helix length, a single-stranded monomer buries one of its termini in the bilayer. Exposure of the carboxyl terminus to the bilayer surface and lack of exposure for the amino terminus was documented by shift reagent NMR experiments (11). It has been observed that the native monovalent cation selective channel function is maintained when formal charges are introduced at the carboxyl termin...

show abstract

mentioning

confidence: 52%

Validation of the single-stranded channel conformation of gramicidin A by solid-state NMR

Kovacs

Quine

Cross

1999

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

View full text Add to dashboard Cite

show abstract

“…Structural (22,52) and functional studies (5) show that the predominant conformation of gA in lipid bilayers is the SS ␤ 6.3 -helical channel structure. The four Trps at the C-terminal cluster at the membrane-water interface (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Implication for Membrane Proteins-Although Trp often is considered to be hydrophobic, Trp residues have long been described as "anchoring" residues in membrane proteins (2,4,5), and as early as 1984, it was found, using [Trp 1 ]gA channels, that there is a significant penalty associated with burying Trp residues in the bilayer core (62). More recently, it was shown that Trps near the C terminus of a membrane-spanning ␣-helix display a distinctly different behavior from those near the N terminus, reflected in large differences in the allowed side chain torsion angles and ring motions, highlighting the directionality of an ␣-helix relative to the bilayer/solution interface (67).…”

Section: Discussionmentioning

confidence: 99%

“…gA is a 15-residue peptide with an alternating L-and D-amino acid sequence: formyl-VGALAVVVWLWLWLW-ethanolamide (the D-amino acids are underlined), which forms monovalent cation-permeable transmembrane channels when two 15-residue subunits join from opposite sides of the membrane (5,21). Each gA subunit has four Trps, which in the singlestranded (SS) ␤-helical channel structure are localized near the membrane/water interface ( Fig.…”

mentioning

confidence: 99%

See 1 more Smart Citation

The Preference of Tryptophan for Membrane Interfaces

Sun

Greathouse

Andersen

et al. 2008

Journal of Biological Chemistry

Self Cite

100

View full text Add to dashboard Cite

To better understand the structural and functional roles of tryptophan at the membrane/water interface in membrane proteins, we examined the structural and functional consequences of Trp 3 1-methyl-tryptophan substitutions in membranespanning gramicidin A channels. Gramicidin A channels are miniproteins that are anchored to the interface by four Trps near the C terminus of each subunit in a membrane-spanning dimer. We masked the hydrogen bonding ability of individual or multiple Trps by 1-methylation of the indole ring and examined the structural and functional changes using circular dichroism spectroscopy, size exclusion chromatography, solid state 2 H NMR spectroscopy, and single channel analysis. N-Methylation causes distinct changes in the subunit conformational preference, channel-forming propensity, single channel conductance and lifetime, and average indole ring orientations within the membrane-spanning channels. The extent of the local ring dynamic wobble does not increase, and may decrease slightly, when the indole NH is replaced by the non-hydrogen-bonding and more bulky and hydrophobic N-CH 3 group. The changes in conformational preference, which are associated with a shift in the distribution of the aromatic residues across the bilayer, are similar to those observed previously with Trp 3 Phe substitutions. We conclude that indole N-H hydrogen bonding is of major importance for the folding of gramicidin channels. The changes in ion permeability, however, are quite different for Trp 3 Phe and Trp 3 1-methyl-tryptophan substitutions, indicating that the indole dipole moment and perhaps also ring size and are important for ion permeation through these channels.

show abstract

“…This conformation, which may span the whole lipid bilayer with a nonchannel structure, is in a metastable state that slowly dissociates, refolding into the 3.3 6.3 -helical conformation. [7][8][9][10][11][12] Progressive substitution of the four Trp residues, located at the C-terminus end, by Phe residues tends to shift the conformational equilibrium from channel-forming monomers to nonconducting dsdimers in DOPC bilayers. [19][20][21] In the monosubstituted analogues of gramicidin A, the destabilization of the monomeric structure in favor of the ds-dimeric one is greater for Trp substitution in the 9 and 13 positions, while it is definitely smaller in the 11 and 15 positions.…”

Section: Introductionmentioning

confidence: 99%

Gramicidin Conducting Dimers in Lipid Bilayers Are Stabilized by Single-File Ionic Flux along Them

2007

View full text Add to dashboard Cite

Gramicidin D was incorporated in a biomimetic membrane consisting of a lipid bilayer tethered to a mercury electrode via a hydrophilic spacer, and its behavior was investigated in aqueous 0.1 M KCl by potential-step chronocoulometry and electrochemical impedance spectroscopy. The impedance spectra, recorded from 0.1 to 1 × 10 5 Hz over a potential range of 0.7 V, were fitted to a series of RC meshes, which were related to the different substructural elements of the biomimetic membrane. These impedance spectra were compared with those obtained by incorporating valinomycin, under otherwise identical conditions. The potential dependence of the stationary currents reported on bilayer lipid membranes by Bamberg and Läuger (Bamberg, E.; Läuger, P. J. Membrane Biol. 1973, 11, 177-194) as well as those extracted from potential-step chronocoulometric measurements was interpreted by relating the increase in gramicidin dimerization to a progressive increase in single-file K + flux along the dimeric channels. An analogous approach was adopted in explaining the difference between the impedance spectra obtained with gramicidin D and those obtained with valinomycin. It is concluded that gramicidin has a low tendency to form dimers in the absence of ionic flux.

show abstract

Kinetics of Gramicidin Channel Formation in Lipid Bilayers: Transmembrane Monomer Association

Cited by 247 publications

References 30 publications

Validation of the single-stranded channel conformation of gramicidin A by solid-state NMR

Validation of the single-stranded channel conformation of gramicidin A by solid-state NMR

The Preference of Tryptophan for Membrane Interfaces

Gramicidin Conducting Dimers in Lipid Bilayers Are Stabilized by Single-File Ionic Flux along Them

Contact Info

Product

Resources

About