Investigation of an unnatural amino acid for use as a resonance Raman probe: Detection limits, solvent and temperature dependence of the νC≡N band of 4-cyanophenylalanine.

Journal of Raman Spectroscopy : JRS
Colin L WeeksT G Spiro

Abstract

The incorporation of unnatural amino acids into proteins that act as spectroscopic probes can be used to study protein structure and function. One such probe is 4-cyanophenylalanine (PheCN), the nitrile group of which has a stretching mode that occurs in a region of the vibrational spectrum that does not contain any modes from the usual components of proteins and the wavenumber is sensitive to the polarity of its environment. In this work we evaluate the potential of UV resonance Raman spectroscopy for monitoring the sensitivity of the νC≡N band of PheCN incorporated into proteins to the protein environment. Measurement of the Raman excitation profile of PheCN showed that considerable resonance enhancement of the Raman signal was obtained using UV excitation and the best signal-to-noise ratios were obtained with excitation wavelengths of 229 and 244 nm. The detection limit for PheCN in proteins was ~10 μM, approximately a hundred-fold lower than the concentrations used in IR studies, which increases the potential applications of PheCN as a vibrational probe. The wavenumber of the PheCN νC≡N band was strongly dependent on the polarity of its environment, when the solvent was changed from H(2)O to THF it decreased by 8 cm(-1). Th...Continue Reading

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Citations

May 18, 2013·Chemical Reviews·Heejae Kim, Minhaeng Cho
Jun 21, 2013·The Journal of Chemical Physics·Justin E Elenewski, John C Hackett
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