The Aromatic Residues Trp and Phe Have Different Effects on the Positioning of a Transmembrane Helix in the Microsomal Membrane

Braun, Paula; Heijne, Gunnar von

doi:10.1021/bi990923a

Cited by 141 publications

(158 citation statements)

References 27 publications

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“…Perturbations primarily associated with the anisotropic chemical shifts may reflect differences in their chemical shift tensors. Perturbations in the first or last turn are likely to reflect the interaction of this helical turn with the membrane interface through amphipathic side chains, anchoring and stabilizing the TM domain in the lipid bilayer (44,45), and hence the helical backbone may be somewhat perturbed at these sites. However, it is clear that the tilt and rotational orientation of the helices remain constant throughout their lengths.…”

Section: Resultsmentioning

confidence: 99%

“…These helices span nearly 30 Å of the POPC/POPG bilayer hydrophobic thickness. Two tryptophan residues at the helix termini (Trp47 on TM1 and Trp73 on TM2) have their indole N-H oriented toward the membrane interfacial regions and serve as anchors (44,57); two other tryptophans (Trp32 and Trp92) form an interhelical stacking interaction. Neighboring polar and charged residues (Ser30 and Gln52 for TM1 and Tyr75 and Arg91 for TM2) form hydrogen bonds with lipid phosphate and carbonyl groups (Fig.…”

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Das

Dai

Hung

et al. 2014

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

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The 93-residue transmembrane protein CrgA in Mycobacterium tuberculosis is a central component of the divisome, a large macromolecular machine responsible for cell division. Through interactions with multiple other components including FtsZ, FtsQ, FtsI (PBPB), PBPA, and CwsA, CrgA facilitates the recruitment of the proteins essential for peptidoglycan synthesis to the divisome and stabilizes the divisome. CrgA is predicted to have two transmembrane helices. Here, the structure of CrgA was determined in a liquid–crystalline lipid bilayer environment by solid-state NMR spectroscopy. Oriented-sample data yielded orientational restraints, whereas magic-angle spinning data yielded interhelical distance restraints. These data define a complete structure for the transmembrane domain and provide rich information on the conformational ensembles of the partially disordered N-terminal region and interhelical loop. The structure of the transmembrane domain was refined using restrained molecular dynamics simulations in an all-atom representation of the same lipid bilayer environment as in the NMR samples. The two transmembrane helices form a left-handed packing arrangement with a crossing angle of 24° at the conserved Gly39 residue. This helix pair exposes other conserved glycine and alanine residues to the fatty acyl environment, which are potential sites for binding CrgA’s partners such as CwsA and FtsQ. This approach combining oriented-sample and magic-angle spinning NMR spectroscopy in native-like lipid bilayers with restrained molecular dynamics simulations represents a powerful tool for structural characterization of not only isolated membrane proteins, but their complexes, such as those that form macromolecular machines.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Das

Dai

Hung

et al. 2014

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

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“…The pair of tryptophans in the pore helix is highly conserved across K þ channels. Tryptophans preferentially reside at membrane interfaces (17)(18)(19). The complex properties known as aromaticity underlie this preference (17).…”

Section: Discussionmentioning

confidence: 99%

Biogenesis of the pore architecture of a voltage-gated potassium channel

Gajewski

Dagcan

Roux

et al. 2011

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

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The pore domain of voltage-gated potassium (Kv) channels consists of transmembrane helices S5 and S6, the turret, the pore helix, the selectivity filter, and the loop preceding S6, with a tertiary reentrant structure between S5 and S6. Using biogenic intermediates, mass tagging (pegylation), and a molecular tape measure, we explored the possibility that the first stages of pore formation occur prior to oligomerization of the transmembrane core. Pegylation of introduced cysteines shows that the pore helix, but not the turret, forms a compact secondary structure in the terminal 20 Å of the ribosomal tunnel. We assessed the tertiary fold of the pore loop in monomeric constructs by determining the relative accessibilities of select cysteines using the kinetics of pegylation. Turret residues are accessible at the extracellular surface. In contrast, pore helix residues are less accessible. All-atom molecular dynamics simulations of a single Kv monomer in a solvated lipid membrane indicate that secondary and tertiary folds are stable over 650 ns. These results are consistent with acquisition of a tertiary reentrant pore architecture at the monomer stage of Kv biogenesis and begin to define a plausible sequence of folding events in the formation of Kv channels.T he pore of a selective potassium channel has evolved to be highly selective and rapidly conducting. These permeation properties derive from the exquisitely precise architecture of the tetrameric channel protein (1). Voltage-gated potassium (Kv) channels have six transmembrane segments, S1-S6 and a cytosolic T1 tetramerization domain. The pore region is composed of S5, S6, and an intervening pore loop consisting of a turret, the pore helix, the selectivity filter, and a pre-S6 loop (Fig. 1). Reentry of the pore loop between S5 and S6 is necessary to position the selectivity filter facing both intra-and extracellular aqueous vestibules. The pore loop is structurally highly conserved among all ion channels. Folding defects in this region will lead to channelopathies. For example, mutations in this region cause long QT syndrome and can be fatal (2, 3). Folding of Kv channels, as with all proteins, begins as early as the birth of the nascent peptide attached to the ribosome and continues during its tenure in the endoplasmic reticulum (ER) (4-7). Yet, we know little regarding the sequence of events responsible for the critical folding of the pore itself, e.g., when are the secondary and tertiary folds that define the structure of the reentrant pore loop acquired?Here, we explore the possibility that pore structure forms early in biogenesis, in the monomer Kv1.3 channel protein. Fig. 1 (dashed circle) delineates the secondary and tertiary architecture probed in this study. Using a cysteine scan of the pore helix and turret region in a nascent (attached to the ribosome) Kv1.3 and a mass-tagging strategy, we demonstrate that compaction of the pore helix occurs in the distal portion of the ribosomal tunnel near the exit port. Using the kinetics of modification of cysteine...

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“…It has been suggested that Trp residues prefer to be positioned at a well-defined site in the lipid headgroups (3,(11)(12)(13)(14)(15) and that thereby they can act as membrane anchors. Their abundance at the lipid/water interface in several membrane proteins (5-7) is therefore likely to be functionally important, e.g.…”

mentioning

confidence: 99%

Interfacial Positioning and Stability of Transmembrane Peptides in Lipid Bilayers Studied by Combining Hydrogen/Deuterium Exchange and Mass Spectrometry

Demmers¹,

Duijn²,

Haverkamp³

et al. 2001

Journal of Biological Chemistry

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Nano-electrospray ionization mass spectrometry (ESI-MS) was used to analyze hydrogen/deuterium (H/D) exchange properties of transmembrane peptides with varying length and composition. Synthetic transmembrane peptides were used with a general acetyl-GW 2 (LA) n LW 2 A-ethanolamine sequence. These peptides were incorporated in large unilamellar vesicles of 1,2-dimyristoyl-sn-glycero-3-phosphocholine. The vesicles were diluted in buffered deuterium oxide, and the H/D exchange after different incubation times was directly analyzed by means of ESI-MS. First, the influence of the length of the hydrophobic Leu-Ala sequence on exchange behavior was investigated. It was shown that longer peptide analogs are more protected from H/D exchange than expected on the basis of their length with respect to bilayer thickness. This is explained by an increased protection from the bilayer environment, because of stretching of the lipid acyl chains and/or tilting of the longer peptides. Next, the role of the flanking tryptophan residues was investigated. The length of the transmembrane part that shows very slow H/D exchange was found to depend on the exact position of the tryptophans in the peptide sequence, suggesting that tryptophan acts as a strong determinant for positioning of proteins at the membrane/water interface. Finally, the influence of putative helix breakers was studied. It was shown that the presence of Pro in the transmembrane segment results in much higher exchange rates as compared with Gly or Leu, suggesting a destabilization of the ␣-helix. Tandem MS measurements suggested that the increased exchange takes place over the entire transmembrane segment. The results show that ESI-MS is a convenient technique to gain detailed insight into properties of peptides in lipid bilayers by monitoring H/D exchange kinetics.The precise manner in which membrane proteins are embedded in a lipid bilayer is essential for their structure and function. Important structural and dynamic features, such as stability of the transmembrane segments or their precise positioning at the lipid/water interface, will be determined not only by intrinsic properties of the transmembrane segments, but also by their interaction with surrounding lipids. A convenient way to gain insight into how the special characteristics of transmembrane segments and their interaction with lipids may influence the behavior of membrane proteins is by studying model systems of artificial transmembrane peptides with desired properties in well defined lipid bilayers. Recently, we have described a new method using nano-ESI-MS 1 (1) to study the properties of transmembrane protein segments in model systems by analyzing the kinetics of hydrogen/deuterium (H/D) exchange. The results showed that various populations of amide hydrogen atoms can be distinguished that are characteristic for different regions of the transmembrane segments. These populations are fast exchanging amide hydrogens located in the peptide termini that are exposed to the aqueous phase, intermediately exchan...

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The Aromatic Residues Trp and Phe Have Different Effects on the Positioning of a Transmembrane Helix in the Microsomal Membrane

Cited by 141 publications

References 27 publications

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Structure of CrgA, a cell division structural and regulatory protein fromMycobacterium tuberculosis, in lipid bilayers

Biogenesis of the pore architecture of a voltage-gated potassium channel

Interfacial Positioning and Stability of Transmembrane Peptides in Lipid Bilayers Studied by Combining Hydrogen/Deuterium Exchange and Mass Spectrometry

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