利用¹³C魔角旋转NMR波谱对向光素LOV1-C57S进行固态Photo-CIDNP效应的磁场依赖性研究. 在4.7 T(相当于200 MHz ¹H频率)磁场中, 黑暗条件下只检测到较宽的蛋白质信号; 但在光照条件下, 观测到强烈的固态Photo-CIDNP效应. 与在2.3 T(相当于100 MHz ¹H频率)磁场中得到的全部为发射谱(负)信号的固态Photo-CIDNP图谱(首次在非光合反应系统中观测到的Photo-CIDNP效应)不同, 在4.7 T磁场中测得的光诱导¹³C NMR谱呈现吸收/发射混合峰型, 这种峰型与文献报道的LOV2的液态Photo-CIDNP波谱的峰型相似. 对比两种磁场中所得谱图发现, 向光素LOV1-C57S的固态Photo-CIDNP效应具有强烈的磁场依赖性, 而且不同的核对磁场的依赖程度差别很大, 这种巨大的差别反映出在被测试的自由基对中的超精细相互作用影响因素发生了很大的改变.

Here we present the magnetic field dependence of the solid-state Photo-CIDNP effect observed in phototropin LOV1-C57S using ¹³C magic-angle spinning (MAS) NMR spectroscopy. Both dark and light spectra were measured at 4.7 T (i.e., 200 MHz ¹H frequency) using a spinning frequency of 8 kHz. An Avance 200 MHz spectrometer equipped with 4-mm MAS probe (Bruker, Karlsruhe, Germany) was used for the ¹³C MAS NMR experiments. The sample was packed into a 4-mm sapphire rotor and inserted into the MAS probe. For a homogeneous sample distribution against the rotor wall, the sample was frozen at a very low spinning frequency of 500 Hz. Freezing was monitored on the proton tuning channel as a shift of ca. 0.1 MHz. The variable temperature unit on the spectrometer was set to 235 K. The spinning frequency was increased to 8 kHz after the sample is completely frozen. This frequency and set temperature were used for all ¹³C MAS NMR measurements. A simple Hahn-echo pulse sequence with two-pulse phase modulation proton-decoupling was used. Continuous illumination was supplied by a 1 kW xenon lamp. The cycle delay was 2 s, and the measurement time was about 12 h. In the spectrum obtained in the dark, standard broad protein responses appear. Under illumination, several strong additional light-induced signals appeared. In contrast to the entirely emissive (negative) peaks in the photo-CIDNP MAS NMR spectra observed at 2.3 T (i.e., 100 MHz ¹H frequency), the first observation of this effect in a nonphotosynthetic system, the light induced ¹³C NMR peaks at 4.7 T show mixed absorptive/emissive enhancement pattern. This pattern is reminiscent of the spectra observed by liquid state photo-CIDNP of a LOV2 sample. The observed solid-state photo-CIDNP effect is strongly magnetic field dependent, and this field-dependence is well distinguished for the various nuclei. This large difference in magnetic field dependence reflects the large variety of hyperfine factors found in this comparable small-sized and asymmetric radical pair.