Experimental Determination of Membrane Protein Secondary Structure Using Vibrational and CD Spectroscopies

Williams, Robert W.

doi:10.1007/978-1-4614-7515-6_8

Cited by 4 publications

(4 citation statements)

References 88 publications

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“…This method works well for ECD to predict helical content, and also well for VCD to get both helix and sheet contributions. ,,, In general, ECD yields the best helix predictions, but amide I′ IR or VCD have better sheet predictions, although adding amide II data to the analyses improves the latter for both helices and sheets. However, combining VCD with ECD or IR spectra (or ECD and IR together) provides the best predictions of fractions for both helix and sheet structures and gains some sensitivity to the contributions of turns and other (disordered) structures. ,, The quality of the predictions was quite dependent on the training set of protein spectra used, both in terms of the number of proteins and the distributions of structural types in the set. Data for proteins in H 2 O solution were shown to give similar analyses using the amide I and II VCD bands, with somewhat more sensitivity (less error) for helices and disorder than obtained with just the amide I′ VCD in D 2 O .…”

Section: Protein Applicationsmentioning

confidence: 99%

“…However, combining VCD with ECD or IR spectra (or ECD and IR together) provides the best predictions of fractions for both helix and sheet structures and gains some sensitivity to the contributions of turns and other (disordered) structures. 619,656,657 The quality of the predictions was quite dependent on the training set of protein spectra used, both in terms of the number of proteins and the distributions of structural types in the set. Data for proteins in H 2 O solution were shown to give similar analyses using the amide I and II VCD bands, with somewhat more sensitivity (less error) for helices and disorder than obtained with just the amide I′ VCD in D 2 O.…”

Section: Secondary Structure Analyses With Vcdmentioning

confidence: 99%

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Structure of Condensed Phase Peptides: Insights from Vibrational Circular Dichroism and Raman Optical Activity Techniques

2020

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Peptides and proteins are naturally chiral molecular systems so that sensing their structure and conformation with chirality-based spectral methods is an obvious and long-used diagnostic application. Extending chiroptical techniques to measurement of vibrational transitions, in the form of vibrational circular dichroism (VCD) and Raman optical activity (ROA), expands the number and types of excitations available that might provide structural insight and can provide an alternate and, in some cases, a more distinctive conformational probe. Since the dominant repeating structural element in peptides is the locally achiral amide group, VCD senses the polymeric structure through amide coupling, which is directly dependent on secondary structure. Determination of the type and relative contribution of these structural components through empirical correlation with spectral character has been the main application of VCD for peptides and proteins, although this is now reinforced by extensive theoretical modeling. Monitoring structural and conformational change induced by environmental perturbations provides another important application. More recently, VCD has been used to detect morphological variations in fibril states of aggregated peptides and proteins. ROA has parallel secondary structural sensitivities, with more applications for proteins than peptides, and has more sensitivity to local configuration and side chains. This review covers the range of peptide studies done with VCD and extends them to compare with example protein and ROA applications.

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Section: Protein Applicationsmentioning

confidence: 99%

Section: Secondary Structure Analyses With Vcdmentioning

confidence: 99%

Structure of Condensed Phase Peptides: Insights from Vibrational Circular Dichroism and Raman Optical Activity Techniques

2020

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“…194,233,235 This toolbox was more recently extended to specifically deal with common problems present in models derived from medium-resolution cryo-EM maps (>3 Å) with the CaBLAM (C-Alpha Based Low-Resolution Annotation Method) tool. 233,236 CaBLAM uses Cα angular distributions to assess main-chain geometry and identify secondary structure. Although the Cα trace is a prominent feature in cryo-EM maps at $3 Å resolution range, the carbonyl bonds of the backbone are not visible, and peptide bond angles are commonly incorrectly orientated.…”

Section: Protein Stereo-chemistry Methodsmentioning

confidence: 99%

Integrating model simulation tools and cryo‐electron microscopy

Beton

Cragnolini

Kaleel

et al. 2022

WIREs Comput Mol Sci

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The power of computer simulations, including machine-learning, has become an inseparable part of scientific analysis of biological data. This has significantly impacted the field of cryogenic electron microscopy (cryo-EM), which has grown dramatically since the "resolution-revolution." Many maps are now solved at 3-4 Å or better resolution, although a significant proportion of maps deposited in the Electron Microscopy Data Bank are still at lower resolution, where the positions of atoms cannot be determined unambiguously. Additionally, cryo-EM maps are often characterized by a varying local resolution, partly due to conformational heterogeneity of the imaged molecule. To address such problems, many computational methods have been developed for cryo-EM map reconstruction and atomistic model building. Here, we review the development in algorithms and tools for building models in cryo-EM maps at different resolutions. We describe methods for model building, including rigid and flexible fitting of known models, model validation, small-molecule fitting, and model visualization. We provide examples of how these methods have been used to elucidate the structure and function of dynamic macromolecular machines.

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“…However, high-resolution spectra require substantial protein concentrations, , and both size limits ,, and line broadening can be a problem. Circular dichroism (CD) reports the fractions of the protein in its secondary conformations, − infrared spectroscopy (IR) provides information on the secondary structure of the protein by detecting the change of the hydrogen-bonding interactions of amide groups within the proteins, , and both methods provided information on protein orientation in vesicles . Molecular dynamics (MD) simulations of membrane proteins is a work in progress, but in principle, MD can report on all the atoms and their locations, given sufficient time, computer power, and accurate force fields. ,,− A variety of chemical modification approaches are available in which side-chain functional groups of specific amino acids are labeled with a probe molecule to map protein structures and detect conformational changes, including those at bilayer interfaces, ,− but none of these chemical approaches target the peptide bond itself.…”

Section: Introductionmentioning

confidence: 99%

Simultaneous Determination of Interfacial Molarities of Amide Bonds, Carboxylate Groups, and Water by Chemical Trapping in Micelles of Amphiphiles Containing Peptide Bond Models

Zhang

Romsted

Zhuang³

et al. 2012

Langmuir

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Chemical trapping is a powerful approach for obtaining experimental estimates of interfacial molarities of weakly basic nucleophiles in the interfacial regions of amphiphile aggregates. Here, we demonstrate that the chemical probe 4-hexadecyl-2,6-dimethylbenzenediazonium ion (16-ArN(2)(+)) reacts competitively with interfacial water, with the amide carbonyl followed by cleavage of the headgroups from the tail at the amide oxygen, and with the terminal carboxylate groups in micelles of two N-acyl amino-acid amphiphiles, sodium N-lauroylsarcosinate (SLS) and sodium N-lauroylglycinate (SLG), simple peptide bond model amphiphiles. Interfacial molarities (in moles per liter of interfacial volume) of these three groups were obtained from product yields, assuming that selectivity toward a particular nucleophile compared to water is the same in an aqueous reference solution and in the interfacial region. Interfacial carboxylate group molarities are ~1.5 M in both SLS and SLG micelles, but the concentration of the amide carbonyl for SLS micelles is ~4.6-5 times less (ca. 0.7 M) than that of SLG micelles (~3 M). The proton on the secondary N of SLG helps solubilize the amide bond in the aqueous region, but the methyl on the tertiary N of SLS helps solubilize the amide bond in the micellar core, reducing its reaction with 16-ArN(2)(+). Application of chemical trapping to proteins in membrane mimetic interfaces should provide insight into the topology of the protein within the interface because trapping of the amide carbonyl and cleavage at the C-N bond occurs only within the interface, and fragment characterization marks those peptide bonds located within the interface.

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Experimental Determination of Membrane Protein Secondary Structure Using Vibrational and CD Spectroscopies

Cited by 4 publications

References 88 publications

Structure of Condensed Phase Peptides: Insights from Vibrational Circular Dichroism and Raman Optical Activity Techniques

Structure of Condensed Phase Peptides: Insights from Vibrational Circular Dichroism and Raman Optical Activity Techniques

Integrating model simulation tools and cryo‐electron microscopy

Simultaneous Determination of Interfacial Molarities of Amide Bonds, Carboxylate Groups, and Water by Chemical Trapping in Micelles of Amphiphiles Containing Peptide Bond Models

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