Mimic of the Green Fluorescent Protein β‑Barrel: Photophysics and Dynamics of Confined Chromophores Defined by a Rigid Porous Scaffold

Chromophores
with a benzylidene imidazolidinone core define the
emission profile of commonly used biomarkers such as the green fluorescent
protein (GFP) and its analogues. In this communication, artificially
engineered porous scaffolds have been shown to mimic the protein β-barrel
structure, maintaining green fluorescence response and conformational
rigidity of GFP-like chromophores. In particular, we demonstrated
that the emission maximum in our artificial scaffolds is similar to
those observed in the spectra of the natural GFP-based systems. To
correlate the fluorescence response with a structure and perform a
comprehensive analysis of the prepared photoluminescent scaffolds, 13C cross-polarization magic angle spinning solid-state (CP-MAS)
NMR spectroscopy, powder and single-crystal X-ray diffraction, and
time-resolved fluorescence spectroscopy were employed. Quadrupolar
spin–echo solid-state 2H NMR spectroscopy, in combination
with theoretical calculations, was implemented to probe low-frequency
vibrational dynamics of the confined chromophores, demonstrating conformational
restrictions imposed on the coordinatively trapped chromophores. Because
of possible tunability of the introduced scaffolds, these studies
could foreshadow utilization of the presented approach toward directing
a fluorescence response in artificial GFP mimics, modulating a protein
microenvironment, and controlling nonradiative pathways through chromophore
dynamics.

以亚苄基咪唑烷酮为核心的生色团，决定了绿色荧光蛋白（green fluorescent protein, GFP）及其类似物等常用生物标志物的发光特性。本研究通讯中，工程化人工多孔支架被证实可模拟蛋白质β桶结构，维持类GFP生色团的绿色荧光响应与构象刚性。尤为关键的是，本研究证实，该人工支架的发射最大值与天然GFP基体系光谱中观测到的结果相近。为将荧光响应与结构建立关联，并对所制备的光致发光支架开展全面分析，本研究采用了13C交叉极化魔角旋转固态（CP-MAS）核磁共振波谱法、粉末与单晶X射线衍射技术，以及时间分辨荧光光谱法。结合理论计算，我们还通过四极自旋回波固态氘核磁共振波谱法，探究了受限生色团的低频振动动力学，证实了配位捕获的生色团所受到的构象限制。鉴于所构建的支架具备可调控性，本研究成果或可为人工GFP模拟体系中定向调控荧光响应、调节蛋白质微环境，以及通过生色团动力学控制非辐射路径提供潜在的应用思路。