Muiño Pedro L, Callis Patrik R
Department of Chemistry, Saint Francis University, Loretto, Pennsylvania 15940, USA.
J Phys Chem B. 2009 Mar 5;113(9):2572-7. doi: 10.1021/jp711513b.
Hybrid quantum mechanical/molecular mechanics (QM-MM) calculations [Callis and Liu, J. Phys. Chem. B 2004, 108, 4248-4259] make a strong case that the large variation in tryptophan (Trp) fluorescence yields in proteins is explained by ring-to-backbone amide electron transfer, as predicted decades ago. Quenching occurs in systems when the charge transfer (CT) state is brought below the fluorescing state (1L(a)) as a result of strong local electric fields. To further test this hypothesis, we have measured the fluorescence quantum yield in solvents of different polarity for the following systems: N-acetyl-L-tryptophanamide (NATA), an analogue for Trp in a protein; N-acetyl-L-tryptophan ethyl ester (NATE), wherein the Trp amide is replaced by an ester group, lowering the CT state energy; and 3-methylindole (3MI), a control wherein this quenching mechanism cannot take place. Experimental yields in water are 0.31, 0.13, and 0.057 for 3MI, NATA, and NATE, respectively, whereas, in the nonpolar aprotic solvent dioxane, all three have quantum yields near 0.35, indicating the absence of electron transfer. In alkyl alcohols the quantum yield for NATA and NATE is between that found for water and that found for dioxane, and it is surprisingly independent of chain length (varying from methanol to decanol), revealing that microscopic H-bonding, and not the bulk dielectric constant, dictates the electron transfer rate. QM-MM calculations indicate that, when averaged over the six rotamers, the greatly increased quenching found in water relative to dioxane can be attributed mainly to the larger fluctuations of the energy gap in water. These experiments and calculations are in complete accord with quenching by a solvent stabilized charge transfer from ring to amide state in proteins.
混合量子力学/分子力学（QM-MM）计算[卡利斯和刘，《物理化学杂志B》2004年，108卷，4248 - 4259页]有力地证明，正如几十年前所预测的那样，蛋白质中色氨酸（Trp）荧光产率的巨大差异是由环到主链酰胺的电子转移所解释的。当由于强局部电场使电荷转移（CT）态低于荧光态（1L(a)）时，系统中就会发生猝灭。为了进一步检验这一假设，我们测量了以下体系在不同极性溶剂中的荧光量子产率：N - 乙酰 - L - 色氨酸酰胺（NATA），蛋白质中Trp的类似物；N - 乙酰 - L - 色氨酸乙酯（NATE），其中Trp酰胺被酯基取代，降低了CT态能量；以及3 - 甲基吲哚（3MI），一个该猝灭机制不会发生的对照物。3MI、NATA和NATE在水中的实验产率分别为0.31、0.13和0.057，而在非极性非质子溶剂二氧六环中，三者的量子产率都接近0.35，表明不存在电子转移。在烷基醇中，NATA和NATE的量子产率介于水中和二氧六环中的产率之间，并且令人惊讶地与链长无关（从甲醇到癸醇变化），这表明微观氢键而非体相介电常数决定了电子转移速率。QM-MM计算表明，当对六种旋转异构体进行平均时，相对于二氧六环，在水中发现的猝灭大大增加主要可归因于水中能隙的较大波动。这些实验和计算与蛋白质中从环到酰胺态的溶剂稳定电荷转移猝灭完全一致。
