A Solid State ¹³C NMR, Crystallographic, and Quantum Chemical Investigation of Phenylalanine and Tyrosine Residues in Dipeptides and Proteins

We report the results of a solid-state NMR and quantum chemical investigation of the 13Cγ NMR chemical shifts in phenylalanine and tyrosine in dipeptides and proteins. Accurate computation of the experimental shifts is shown to require a good description of local electrostatic field effects, and we find the best results (R2 = 0.94, rmsd = 1.6 ppm, range = 17.1 ppm, N = 14) by using a self-consistent reaction field continuum model. There are no obvious correlations with φ, ψ, χ1, or χ2 torsion angles, unlike the results seen with other amino acids. There is, however, a linear relation between computed Cγ atomic charges and shifts for the 14 peptide as well as 18 protein residues investigated. This result is similar to the correlation reported in the 1960s between π-electron density and 13C shifts for classical 4n + 2 (n = 0, 1, 2) π-electron aromatic species, such as cyclopentadienide and the tropylium cation, and in fact, we found that the shielding/atomic charge correlation seen in the peptides and proteins is virtually identical to that seen with a broad range of aromatic carbocations/carbanions. These results suggest the dominance of an electrostatic field polarization model in which increasing π electron density results in an increase in Cγ atomic charge and increased shielding (of σ11 and σ22, perpendicular to the π orbital) in Phe and Tyr, as well as in the other aromatic species. These results are of general interest since they demonstrate the importance of electrostatic field effects on Phe and Tyr Cγ chemical shifts in peptides and proteins and imply that inclusion of these effects will be necessary in order to interpret the shifts of other aromatic species, such as drug molecules, bound to proteins.本研究报道了二肽与蛋白质中苯丙氨酸（phenylalanine, Phe）和酪氨酸（tyrosine, Tyr）的13Cγ核磁共振化学位移的固体核磁共振（Solid-State NMR）与量子化学研究结果。精准计算实验化学位移需对局部静电场效应进行充分描述；本研究采用自洽反应场连续介质模型（self-consistent reaction field continuum model）获得了最优拟合结果（决定系数R²=0.94，均方根偏差rmsd=1.6 ppm，数值范围17.1 ppm，样本量N=14）。与其他氨基酸的相关研究结果不同，本研究未观测到苯丙氨酸与酪氨酸的Cγ化学位移与φ、ψ、χ1或χ2二面角存在显著相关性。但在所研究的14个二肽体系与18个蛋白质残基中，计算得到的Cγ原子电荷与化学位移呈线性相关关系。该线性相关性与20世纪60年代报道的经典4n+2（n=0、1、2）π电子芳香物种的π电子密度与13C化学位移间的相关性一致；进一步发现，二肽与蛋白质体系中的屏蔽效应/原子电荷相关性，与大量芳香碳正离子（carbocations）、碳负离子（carbanions）体系中的相关性几乎完全一致。上述结果表明，静电场极化模型占据主导地位：π电子密度升高会使苯丙氨酸、酪氨酸以及其他芳香物种的Cγ原子电荷增加，同时提升其垂直于π轨道的σ11与σ22分量的屏蔽效应。本研究结果具有广泛参考价值：其证实了静电场效应对二肽与蛋白质中苯丙氨酸、酪氨酸Cγ化学位移的重要性，同时提示，若要解析结合于蛋白质的其他芳香物种（如药物分子）的化学位移，纳入此类静电场效应是必要的。