Origins of the Sensitivity of Molecular Vibrations to Electric Fields: Carbonyl and Nitrosyl Stretches in Model Compounds and Proteins

The vibrational Stark effect (VSE) is the effect of an electric field on the vibrational spectrum. Typically, the largest change induced by an electric field is the linear frequency shift whose sensitivity is known as theStark tuning rate Δμ; the magnitude and direction of Δμ are measured by VSE spectroscopy. This can be used to calibrate the sensitivity of vibrational frequencies to local electrostatic fields in proteins. To be most useful, it is critical to obtain a sensitive probe that has a large Δμ, and understanding the origins of Δμ is required. The Stark tuning rate arises from two distinct origins: mechanical anharmonicity (Δμ a n h ) and the change of the bond force constant because of the electronic polarizability (Δμ b o n d ), with Δμ = Δμ a n h + Δμ b o n d . These two origins were quantitatively investigated for carbonyl and nitrosyl stretches in model compounds and proteins by IR and VSE spectroscopy. The contribution of Δμ a n h was found to dominate Δμ for carbonyl stretches in acetone, methyl vinyl ketone, and 1-methyl-2-pyrrolidinone despite their different degrees of electron delocalization. In contrast, the presence of d-π * back-bonding when CO is bound to the heme causes Δμ b o n d to be more significant, giving rise to large variations in the Fe-CO and C-O stretch frequencies in heme protein variants such as myoglobin mutants, with a strong negative correlation between v F e - C O and v C - O . The back-bonding also increases Δμ a n h because of the enhanced effective charges on the CO oscillator. By comparison, the small variation in Fe-NO stretch frequency and the absence of a strong negative correlation between Fe-NO and N-O stretch frequencies for NO bound to the heme iron in myoglobin variants suggest that Δμ F e - N O is much smaller than Δμ F e C O , possibly because of less back-bonding in heme-NO complexes. Nevertheless, Δμ N - O has approximately the same magnitude as Δμ C - O when bound to heme, suggesting that the anharmonic effect on VSE is larger for the N-O stretch than that for C-O stretch.