Optimized and far-red-emitting variants of fluorescent protein eqFP611
Optimized and far-red-emitting variants of fluorescent protein eqFP611
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DOI：
10.1016/j.chembiol.2008.02.008
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发表时间：
2008-03-01
影响因子：
--
通讯作者：
Wiedenmannl, Joerg
中科院分区：
文献类型：
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作者：
Kredel, Simone;Nienhaus, Karin;Wiedenmannl, Joerg
在光谱的远红区域发射的荧光蛋白（FP）对于全身成像应用是非常有利的，因为长波长光的散射和吸收在组织中显著减少。我们表征了红色荧光蛋白eqFP 611的变体，其具有高达639 nm的明亮荧光发射。额外的红移是由发色团的反-顺异构化引起的。反式和顺式构象之间的平衡受到氨基酸残基143和158的强烈影响。通过进一步的基因工程获得假单体标签。对于eqFP 611变体的红色发色团，通过不受具有非功能性红色发色团的分子的存在影响的方法测定了高达类似于150，000的摩尔消光系数。明亮的荧光使得红移的eqFP 611变体有希望成为开发近红外荧光标记物的先导结构。红色荧光蛋白在细胞生物学应用中表现良好，包括双光子成像。
Fluorescent proteins (FPs) emitting in the far-red region of the spectrum are highly advantageous for whole-body imaging applications because scattering and absorption of long-wavelength light is markedly reduced in tissue. We characterized variants of the red fluorescent protein eqFP611 with bright fluorescence emission shifted up to 639 nm. The additional red shift is caused by a trans-cis isomerization of the chromophore. The equilibrium between the trans and cis conformations is strongly influenced by amino acid residues 143 and 158. Pseudo monomeric tags were obtained by further genetic engineering. For the red chromophores of eqFP611 variants, molar extinction coefficients of up to similar to 150,000 were determined by an approach that is not affected by the presence of molecules with nonfunctional red chromophores. The bright fluorescence makes the red-shifted eqFP611 variants promising lead structures for the development of near-infrared fluorescent markers. The red fluorescent proteins performed well in cell biological applications, including two-photon imaging.