Petchprayoon Chutima, Marriott Gerard
Department of Bioengineering, University of California, Berkeley, CA 94720.
Curr Protoc Chem Biol. 2010 Aug 1;2(3):153-169. doi: 10.1002/9780470559277.ch100054.
Optical switches represent a new class of molecular probe with applications in high contrast imaging and optical manipulation of protein interactions. Small molecule, organic optical switches based on nitrospirobenzopyran (NitroBIPS) and their reactive derivatives and conjugates undergo efficient, rapid and reversible, orthogonal optically-driven transitions between a colorless spiro (SP) state and a colored merocyanine (MC) state. The excited MC-state also emits fluorescence, which serves as readout of the state of the switch. Defined optical perturbations of SP and MC generate a defined waveform of MC-fluorescence that can be isolated against unmodulated background signals by using a digital optical lock-in detection approach or to control specific dipolar interactions on proteins. The protocols describe general procedures for the synthesis and spectroscopic characterization of NitroBIPS and specifically labeled conjugates along with methods for the manipulation of dipolar interactions on proteins and imaging of the MC-state of NitroBIPS within living cells.
光开关代表了一类新型分子探针，可应用于高对比度成像以及蛋白质相互作用的光学操纵。基于硝基螺苯并吡喃（NitroBIPS）的小分子有机光开关及其反应性衍生物和共轭物，在无色螺环（SP）状态和有色部花青（MC）状态之间经历高效、快速且可逆的正交光驱动转变。激发态的MC状态还会发出荧光，可作为开关状态的读出信号。对SP和MC进行特定的光扰动会产生特定波形的MC荧光，通过使用数字光学锁相检测方法，可将其与未调制的背景信号区分开来，或者用于控制蛋白质上特定的偶极相互作用。这些方案描述了NitroBIPS及其特异性标记共轭物的合成和光谱表征的一般程序，以及操纵蛋白质上偶极相互作用和在活细胞内对NitroBIPS的MC状态进行成像的方法。
