Tryptophan Raman bands sensitive to hydrogen bonding and side‐chain conformation

Miura, Tadao; Takeuchi, Hideo; Harada, Issei

doi:10.1002/jrs.1250201007

Cited by 263 publications

(407 citation statements)

References 14 publications

(1 reference statement)

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“…In intact viruses, this Trp signal is a single peak centered at 877 cm Ϫ1 , and after heat treatments, the signal divides to two peaks at 873 and 881 cm Ϫ1 , indicating changes in Trp hydrogen bonding status. The frequency of the Trp signal near 1,550 cm Ϫ1 has been reported to be indicative of changes in side chain conformation (46)(47)(48), and there are heat-induced changes in this region in our data (Fig. 5D).…”

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confidence: 71%

Raman Spectroscopic Signatures of Echovirus 1 Uncoating

Ruokola

Dadu

Kazmertsuk

et al. 2014

J Virol

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In recent decades, Raman spectroscopy has entered the biological and medical fields. It enables nondestructive analysis of structural details at the molecular level and has been used to study viruses and their constituents. Here, we used Raman spectroscopy to study echovirus 1 (EV1), a small, nonenveloped human pathogen, in two different uncoating states induced by heat treatments. Raman signals of capsid proteins and RNA genome were observed from the intact virus, the uncoating intermediate, and disrupted virions. Transmission electron microscopy data revealed general structural changes between the studied particles. Compared to spectral characteristics of proteins in the intact virion, those of the proteins of the heat-treated particles indicated reduced ␣-helix content with respect to ␤-sheets and coil structures. Changes observed in tryptophan and tyrosine signals suggest an increasingly hydrophilic environment around these residues. RNA signals revealed a change in the environment of the genome and in its conformation. The ionized-carbonyl vibrations showed small changes between the intact virion and the uncoating intermediate, which points to cleavage of salt bridges in the protein structure during the uncoating process. In conclusion, our data reveal distinguishable Raman signatures of the intact, intermediate, and disrupted EV1 particles. These changes indicate structural, chemical, and solute-solvent alterations in the genome and in the capsid proteins and lay the essential groundwork for investigating the uncoating of EV1 and related viruses in real time. IMPORTANCEIn order to combat virus infection, we need to know the details of virus uncoating. We present here the novel Raman signatures for opened and intact echovirus 1. This gives hope that the signatures may be used in the near future to evaluate the ambient conditions in endosomes leading to virus uncoating using, e.g., coherent anti-Stokes Raman spectroscopy (CARS) imaging. These studies will complement structural studies on virus uncoating. In addition, Raman/CARS imaging offers the possibility of making dynamic live measurements in vitro and in cells which are impossible to measure by, for example, cryo-electron tomography. Furthermore, as viral Raman spectra can be overwhelmed with various contaminants, our study is highly relevant in demonstrating the importance of sample preparation for Raman spectroscopy in the field of virology. Picornaviruses are small, nonenveloped human pathogens able to cause a wide range of illnesses (1). The subgroup enteroviruses include clinically important echoviruses, coxsackieviruses, and poliovirus. These viruses cause a variety of diseases varying from mild infections to aseptic meningitis, heart muscle damage, and paralysis. Enteroviruses have also recently been associated with chronic diseases such as type 1 diabetes, cardiomyopathies, and atherosclerosis (2, 3). Structural details of these viruses have been obtained by means of X-ray crystallography or at a lower resolution by cryo-electron microsc...

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supporting

confidence: 71%

Raman Spectroscopic Signatures of Echovirus 1 Uncoating

Ruokola

Dadu

Kazmertsuk

et al. 2014

J Virol

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“…The indole ring vibration W3 changes in frequency as a function of the torsional angle 2,1 of the C δ1 -C γ -C -C R linkage (27). As | 2,1 | increases between 60°and 120°, the W3 frequency increases in the range of 1542-1557 cm -1 .…”

Section: A Comparison Of the Solution Structures Of Im7* And Im9* By mentioning

confidence: 99%

Ultraviolet Resonance Raman Studies Reveal the Environment of Tryptophan and Tyrosine Residues in the Native and Partially Folded States of the E Colicin-Binding Immunity Protein Im7

et al. 2005

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Understanding the nature of partially folded proteins is a challenging task that is best accomplished when several techniques are applied in combination. Here we present ultraviolet resonance Raman (UVRR) spectroscopy studies of the E colicin-binding immunity proteins, Im7* and Im9*, together with a series of variants of Im7* that are designed to trap a partially folded state at equilibrium. We show that the environments of the tryptophan and tyrosine residues in native wild-type Im7* and Im9* are indistinguishable, in contrast with models for their structures based on X-ray and NMR methods. In addition, we show that there is a general increase in the hydrophobicity in the environment of Trp75 in all of the variants compared with wild-type Im7*. These data suggest that a significant rearrangement of the tryptophan pocket occurs in the variants, which, together with an overall decrease in solvent accessibility of Trp75 as judged by time-resolved fluorescence lifetime measurements and fluorescence quenching experiments, rationalize the unusual fluorescence properties of the variants reported previously. The data highlight the power of UVRR in analyzing the structural properties of different conformational states of the same protein and reveal new information about the structural rearrangements occurring during Im7* folding, not possible using other spectroscopic methods alone. Finally, we describe a previously unreported dependence of the tryptophan Fermi doublet on excitation wavelength in the ultraviolet region revealed by these protein spectra. We corroborated this observation using tryptophan-containing model compounds and conclude that the conventional interpretation of this UVRR feature at these wavelengths is unreliable.The biological role of the E colicin-binding immunity proteins Im2, Im7, Im8, and Im9 is to protect the host cell from the toxic endonuclease activity of a coexpressed cognate colicin, by binding to an exosite in the DNase domain, inhibiting access of the substrate DNA to the active site of the colicin by steric hindrance and electrostatic repulsion (1, 2). The small size of the immunity proteins (85-87 amino acid residues), their high level of sequence identity (approximately 60%), the single conserved tryptophan, and their lack of disulfide bonds, prosthetic groups, and cis-prolines in the native state make the immunity protein family ideal for protein folding studies. Despite the high degree of similarity between Im7 and Im9 in both sequence and structure, these proteins fold by mechanisms with different levels of kinetic complexity at neutral pH, suggesting that local sequence variations, rather than topology or global stability, are responsible for the observed folding kinetics of these proteins (3-7). Im7* 1 and Im9* (hexahistidinetagged analogues of Im7 and Im9) contain four helices ( Figure 1A), each of which displays polar side chains on the surface of the protein, while the majority of hydrophobic side chains are buried, forming a well-packed hydrophobic core. Im7* and Im...

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“…Direct correlation between the Raman frequency of W3 mode and the absolute value of the torsional angle  2,1 is known [48]. This question can be answered by off-diagonal peaks, so called cross peaks.…”

Section: Tablesmentioning

confidence: 99%