【JACS】告别传统酸碱剂！首个水溶性光控分子开关问世，基本pH区实现超1.7单位精准逆转
文章标题：A Water-Soluble Diarylethene Base for Light-Controlled pH Modulation in Biological Systems
通讯作者：Javier Read de Alaniz
加州大学圣芭芭拉分校的Javier Read de Alaniz教授团队成功开发出首个高水溶性、具备双向调节功能的氮杂环亚胺二芳基乙烯（NHI-DAE）光控碱分子开关。该分子在水中的溶解度超过50 mM，能够通过紫外光和可见光的交替照射，在生物学关键的偏碱性区域内实现大于1.7个pH单位的快速、可逆平衡调节，并成功应用于控制反射蛋白质的动态自组装。
引言
在生物和化学系统中，空间的酸碱度（pH）精密调控对酶催化、蛋白质折叠及信号传导至关重要。虽然科学家此前开发出多种“光致产酸剂”或“光致产碱剂”，但它们大多不可逆，或者在水中的溶解度极低（通常小于300 M），必须依赖有机共溶剂，这极易破坏生物大分子的天然活性。因此，开发一种能够在纯水相、特别是中性至碱性生物环境中表现出高溶解度、大pH变化范围且高度可逆的光控开关，一直是化学与合成生物学领域的重大挑战。
Figure 1. (A) Merocyanine photoacids are water-soluble in millimolar-scale concentrations and reversibly switch pH in the acidic and neutral pH regimes. In this work, a water-soluble photoswitch that reversibly switches pH in the neutral-basic regime is presented. (B) Structure of the N-heterocyclic imine diarylethene (NHI-DAE) photoswitchable base prepared in this study. The open diarylethene isomer is strongly basic and will undergo electrocyclization to the weakly basic, closed isomer with 300 nm light. The cycloreversion reaction is triggered by >500 nm.
主要实验及结论
为了打破溶解度瓶颈，研究人员通过精妙的化学修饰，在如图1所示的氮杂环亚胺（NHI）骨架上引入了具有空间位阻的季铵盐亲水基团。这一改进使得全新的NHI-DAE分子开关在纯水中的溶解度骤增至50 mM以上（如图2所示）。光谱实验表明，该分子在300 nm紫外光照射下会发生电环化反应，转变为紫色闭环结构；而在大于500 nm的可见光照射下则完全恢复。
Figure 2. (A) Synthesis of NHI-DAE photoswitchable base. (B) Synthesized photoswitch is soluble to >50 mM in Milli-Q water.
研究团队通过光谱滴定深入解析了其光控酸碱机理。如图3所示，紫外光引发的闭环反应破坏了原有的芳香稳定化结构，导致其亚胺氮原子的解离常数从9.6大幅下降至7.9，表现出明显的“光致产酸”行为；而在低pH区间，由于环化消除了特定空间位阻，端基氮原子的basic性反而增强，表现出“光致产碱”的特质。如图4所示的模型预测与实验结果高度吻合，在3.5 mM的无缓冲体系中，紫外光仅需照射3分钟，溶液pH值便剧烈下降1.70个单位，且该过程可通过白光照射轻松逆转，并成功稳定循环10次以上。
Figure 3. (A) UV–vis absorbance spectra of 14.4 μM solution of open-NHI-DAE in a 20 mM sodium phosphate buffer over a range of pH values. (B) UV–vis absorbance spectra of 14.4 μM solution of closed-NHI-DAE in a 20 mM sodium phosphate buffer over a range of pH values. (C) Schematic of the four protonation states of the NHI-DAE photoswitch. Counterions omitted for simplicity. (D) Photoresponsive change in p_K_a between the open and closed isomers. (E) Normalized model of spectrophotometric p_K_a titration curve for open-NHI-DAE and closed-NHI-DAE at λ = 224 nm, created with experimentally determined p_K_a values.
Figure 4. Photo-pH properties of a 3.5 mM NHI-DAE solution with 20 mM NaCl in Milli-Q water. (A) Model predicting pH changes of a 3.5 mM NHI-DAE solution under 300 nm irradiation. Compared to experimental pH changes under 300 nm irradiation. (B) The pH of the DAE solution can be switched at least 10 times with 300 nm light and white light (400 nm ≤ λ ≤ 800 nm).
最后，团队将这一光控开关应用于仿生光学领域。如图5所示，他们利用NHI-DAE诱导的pH剧烈变化，成功调控了源自乌贼的阳离子反射蛋白质（Reflectin）的电荷中和与自组装行为。在光控pH驱动下，反射蛋白溶液的浊度随组装体尺寸变化产生可逆波动，透射电镜（TEM）也证实其平均微米级粒径在光照后发生了显著缩减。
Figure 5. (A) Schematic diagram illustrating the pH-responsive assembly behavior of the wild-type and 6E reflectin mutants. In this cartoon, open-NHI-DAE and closed-NHI-DAE are represented as the white and purple molecules, respectively. At a higher pH, more of reflectin’s positive charges are neutralized which drives formation of larger protein assemblies. When the pH decreases under 300 nm light irradiation, the sizes of the protein assemblies decrease, which reduces macroscopic solution turbidity, as measured via UV–vis spectroscopy. (B) UV–vis spectra of a 3.5 mM DAE, 4 μM reflectin 6E, and 20 mM NaCl solution before (gray trace) and after (purple trace) irradiation with 300 nm light. The raised baseline for λ ≥ 700 nm is attributed to turbidity. (C) Turbidity of the reflectin 6E-DAE samples, measured by absorbance at 800 nm, is cyclable with UV and white light irradiation. The sizes of reflectin 6E assemblies were measured using transmission electron microscopy (TEM) (D) before (initial pH = 9.6) and (E) after irradiation with 300 nm light. Representative TEM images are shown with a scale bar of 2 μm.
总结及展望
该研究成功构建了兼具高水溶性、宽响应窗口和大pH动态变化范围的光控分子开关。这种创新的分子设计不仅突破了有机溶剂对生物体系的限制，更在偏碱性区间填补了国际光控酸碱技术的空白。未来的研究将进一步聚焦于红外或近红外光激活技术的开发，以期在活体生物成像、智能药物递送以及动态响应功能材料等前沿领域实现更深层次的应用。