Synthesis of 5‐Cyano‐Tryptophan as a Two‐Dimensional Infrared Spectroscopic Reporter of Structure

Abstract

A concise synthesis of protected 5-cyano-L-tryptophan (Trp5CN) has been developed for 2D IR spectroscopic investigations within peptides or proteins. To assess the potential of differently substituted cyano-tryptophans, several model cyanoindole systems were characterized using IR spectroscopy. Upon assessment of their spectroscopic properties, Trp5CN was integrated into a model peptide sequence, Trp5CN–Gly–Phe4CN, to elucidate its structure. This peptide demonstrates the capability of this probe, Trp5CN and Phe4CN, to capture structural information via 2D IR spectroscopy. The 2D IR spectrum of the peptide in water was simulated to reveal a unique spectral signature resulting from the presence of dipolar coupling. The coupling strength between cyano-labels was determined to be 1.4 cm−1 by matching the slopes along the max contour between the simulated and experimental spectrum. Using transition dipole coupling, a distance between the two probes of 13 Å was calculated.

Probing Intermolecular Interactions of Amyloidogenic Fragments of SOD1 by Site-Specific Tryptophan and Its Noncanonical Derivative

“…Taken together, those two factors make Trp 5CN a much more sensitive CD probe. Third, Trp 5CN is also an IR probe as the vibrational stretch of the CN group can sense the electrostatic field − and hydration status of the surrounding environment, such as being exposed to solvent or buried in a hydrophobic interior, which is reflected in the frequency and the line width of its FTIR spectra. Caution needs to be exercised because the installation of noncanonical amino acids may perturb amyloid aggregation pathways.…”

Tuning Molecular Vibrational Energy Flow within an Aromatic Scaffold via Anharmonic Coupling

“…Our group, as shown in previous work, has an interest in studying the vibrational dynamics of the azido (N 3 )- and cyano (CN)-transitions within biological systems. ,, These modes have been established as valuable infrared probes due to their sensitivity to their local environment, small size to limit perturbation to the molecular system, and the large transition dipole strength of azides, which increases the signal-to-noise ratio. Finally, in biological systems, these transitions are spectrally isolated from other congested regions of the IR spectrum. ,,, Recently, these transitions have been placed in tandem on molecular systems revealing dipolar coupling for structural studies within different biological scaffolds. , To further examine the coupling mechanisms of these probe pairs, several molecular models were designed and synthesized, including para -azidobenzonitrile (PAB) and para -(azidomethyl)­benzonitrile (PAMB). Within these testing manifolds, our 2D IR measurements detected the presence of some dipole–dipole coupling and extensive IVR.…”

“…For the past two decades, two-dimensional infrared (2D IR) spectroscopy has become the premiere ultrafast technique to characterize vibrational dynamics. It has been used to study a wide array of systems, such as structural dynamics in biomolecules, energy transfer in materials, population exchange, and properties of ion channels in cell membranes. − Additionally, the localized solvent dynamics in heterogeneous solutions, which play an important role in reactivity, are also determined through spectral diffusion observed using 2D IR spectroscopy. − Another valuable characteristic of 2D IR is the ability to detect interactions between multiple vibrational transitions within a molecular system. For example, the presence of vibrational coupling, i.e., mechanical and/or dipole–dipole coupling, ,, is oftentimes monitored through the presence of cross peaks in the 2D IR spectrum at the earliest evolution or waiting times. ,,− Cross peaks also can appear as a function of later waiting times.…”

Synthesis and application of the blue fluorescent amino acid l-4-cyanotryptophan to assess peptide–membrane interactions

“…Therefore, there has been a continuous effort to develop new biological fluorescence reporters that can meet different requirements and goals. For example, over the past few decades many analogues of tryptophan (Trp), which is the most fluorescent native amino acid in proteins, have been examined and explored, [4][5][6][7][8][9][10][11][12][13][14][15][16] for the purpose of identifying Trp-based, unnatural amino acid (UAA) fluorophores that have improved photophyiscal properties over Trp. While Trp is a useful and convenient intrinsic fluorescence probe of protein structure and dynamics, its application is limited to in vitro spectroscopic studies.…”