Syntheses of Highly Fluorescent GFP-Chromophore Analogues

Eight B-containing compounds, i.e., 1a−h, were prepared as mimics of the green fluorescent
protein (GFP) fluorophore. The underlying concept was that synthetic GFP chromophore analogues are
not fluorescent primarily because of free rotation about an aryl−alkene bond (Figure b). This rotation is
not possible in the β-barrel of GFP; hence, the molecule is strongly fluorescent. In compounds 1a−h,
radiationless decay via this mechanism is prevented by complexation of the BF2 entity. The target materials
were prepared via two methods; most were obtained according to the novel route shown in Scheme b,
but compound 1f was made via the procedure described in Scheme . Both syntheses involved formation
of undesired compounds E-4a−h that formed simultaneously with the desired isomeric intermediates Z-4a−h. Both compounds form BF2 adducts, i.e., 1a−h and 5a−h, respectively. Methods used for spectroscopic
characterization and differentiation of compounds in the series 1 and 5 are discussed, and these are
supported by single-crystal X-ray diffraction analyses for compounds 1c, 5c, 1f, and 5f. Electronic spectra
of compounds 1a−h and 5a−h were studied in detail. Those in the 5 series were shown to be only weakly
fluorescent, but the 1 series were strongly fluorescent compounds (comparable to the boraindacene,
BODIPY, dyes). Compounds 1g and 1h are water soluble, and 1h has particularly significant potential as
a probe, since it also has a carboxylic acid group for attachment to biomolecules.

本研究合成了8种含硼化合物（1a−h），作为绿色荧光蛋白（green fluorescent protein, GFP）荧光团的模拟物。本研究的核心设计思路为：合成的GFP荧光团类似物本身荧光极弱，主要原因是其芳基-烯烃键可发生自由旋转（图b）。而在GFP的β桶状结构中，此类旋转受到限制，因此该分子可呈现出强烈的荧光。在1a−h这类化合物中，通过BF₂基团的络合作用，可阻断上述通过旋转实现的无辐射衰变路径。目标化合物通过两种方法制备：多数产物可通过方案b中展示的全新合成路线获得，但化合物1f需依照方案中描述的步骤合成。两种合成路径均会同时生成副产物E-4a−h与目标异构中间体Z-4a−h。两类化合物均可形成BF₂加合物，分别对应1a−h与5a−h。本文讨论了用于1与5系列化合物的光谱表征及结构区分方法，并通过单晶X射线衍射分析，对1c、5c、1f及5f四种化合物进行了结构确证。本研究对1a−h与5a−h系列化合物的电子光谱开展了详细研究。结果表明，5系列化合物仅表现出微弱的荧光，而1系列化合物则具有强烈的荧光性能，其荧光强度可与硼二吡咯亚甲基（boraindacene, BODIPY）类染料相媲美。化合物1g与1h具备水溶性，其中1h更具备作为荧光探针的巨大应用潜力，因其还带有可用于连接生物分子的羧酸基团。