Cyanamide as an Infrared Reporter: Comparison of Vibrational Properties between Nitriles Bonded to N and C Atoms

Abstract

Infrared (IR) probes based on terminally blocked β-cyanamidoalanine (AlaNHCN) 1 and p-cyanamidophenylalanine (PheNHCN) 2 were synthesized, and the vibrational properties of their CN stretch modes were studied using Fourier transform infrared (FTIR) and femtosecond IR pump-probe spectroscopies in combination with quantum chemical calculations. From FTIR studies, it is found that the transition dipole strengths of the cyanamide (NHCN) group in 1 and 2 are much larger than those of the nitrile (CN) group but comparable to those of the isonitrile (NC) and azido (N) groups in their previously studied analogs. The CN stretch frequencies in 1 and 2 are red-shifted from those in their nitrile analogs but more blue-shifted from the NC and N stretch frequencies in their isonitrile and azido analogs. The much larger transition dipole strength and the red-shifted frequency of the cyanamide relative to nitrile group originates from the n → π* interaction between the N atom's nonbonding (n) and CN group's antibonding (π*) orbitals of the NHCN group. Unlike aliphatic cyanamide 1, aromatic cyanamide 2 shows a complicated line shape of the CN stretch spectra. Such a complicated line shape arises from the Fermi resonance between the CN stretch mode of the NHCN group and one of the overtones of the phenyl ring vibrations and can be substantially simplified by deuteration of the NHCN into NDCN group. From IR pump-probe experiments, the vibrational lifetimes of the CN stretch mode in 1 were determined to be 0.58 ± 0.04 ps in DO and 0.89 ± 0.09 ps in HO and those in 2 were determined to be 1.64 ± 0.13 ps in CHOD/dimethyl sulfoxide and 0.30 ± 0.05 and 2.62 ± 0.26 ps in CHOH. The short time component (0.30 ± 0.05 ps) observed for 2 in CHOH is attributed to the vibrational relaxation through Fermi resonance. These vibrational lifetimes are close to those of the nitrile and azido groups but shorter than those of the isonitrile group. Consequently, cyanamide behaves like an apparent vibrational hybrid of nitrile and isonitrile in that cyanamide is similar to nitrile in vibrational frequency and lifetime but to isonitrile in transition dipole strength. It is believed that cyanamide has the potential to be a strongly absorbing IR reporter of the conformational and environmental structure and dynamics of biomolecules in comparison to nitrile, a weak absorber.

Substituent Effects on the Vibrational Properties of the CN Stretch Mode of Aromatic Nitriles: IR Probes Useful for Time‐resolved IR Spectroscopy

“…This means that the larger E (2) is the stronger interaction between the electron donor and the acceptor is. For example, in the case of the CO binding heme, the interaction ( d ‐ π * back bonding) between the d bonding orbital (donor) of the Fe atom and the π * antibonding orbital (acceptor) of the CO group results in weakening the CO bond strength, thereby the increase of the molar extinction coefficient of the CO stretch mode [24] . In a similar way, investigating the interaction between the electron donor and acceptor in CN moiety can help to understand the correlation among the variance of vibrational frequency, bond length, and transition dipole strength of CN stretch mode with the electron‐donating ability of p ‐substituent.…”

“…On the other hand, aliphatic nitrile exhibits small transition dipole strength, making it difficult to be used as an IR probe, although the CN can be introduced at various sites. In our previous study, we reported that cyanamide (NHCN) group has the potential to be a strongly absorbing IR reporter and compared vibrational properties of aliphatic (cyanamide‐derivatized alanine) and aromatic (cyanamide‐derivatized phenylalanine) compounds [24] . Through FT‐IR measurement, we found that transition dipole strengths (in units of 10 −2 D 2 ) of both compounds in dimethylformamide (DMF) solvent (3.78 and 2.33, respectively) are much larger than the counterparts of nitrile (CN) group in the same solvent (0.15 and 0.99, respectively) [25,26] .…”

Azidohomoalanine: A Minimally Invasive, Versatile, and Sensitive Infrared Label in Proteins To Study Ligand Binding

“…[15][16][17][18][19] Such studies require a special vibrational label that absorbs in the spectral window between ≈1700 cm −1 to ≈2800 cm −1 in order to discriminate it from a huge protein background. [20][21][22][23][24][25] Among the possible molecular groups that have been proposed in that regard, 16,17,19,[26][27][28][29][30][31][32][33][34][35][36][37][38][39][40][41][42][43][44] we favor the noncanonical amino acid azidohomoalanine (Aha). Aha has a relatively high extinction coefficient (≈300-400 M −1 cm −1 ), 15 and it has been shown to be an environment-sensitive infrared probe, capable of sensing the polarity of its environment via the frequency of its vibrational transition 15,18,28,[45][46][47] Additionally, this noncanonical amino acid can be incorporated into a protein in virtually any position via methionine-auxotrophic expression strategy.…”

Isonitrile-Derivatized Indole as an Infrared Probe for Hydrogen-Bonding Environments

“…Unfortunately, the application of the azido probe has also been limited owing to its broad bandwidth, short vibrational lifetime, and accidental Fermi resonance [13,14,15]. Recently, the cyanamide group has been reported as an optimized version of the nitrile group, owing to its larger transition dipole strength and longer vibrational lifetime [16]. We have also investigated the diazo group as a site-specific IR probe to detect local hydration environments [17].…”

“…Additionally, the low concentration in a biological sample makes the measurement even harder. Tremendous effort has been dedicated to the development of unnatural amino acid IR probes with greater transition dipole strength [16,17].…”