Brustad Eric M, Lemke Edward A, Schultz Peter G, Deniz Ashok A
Department of Molecular Biology, Department of Chemistry and the Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
J Am Chem Soc. 2008 Dec 31;130(52):17664-5. doi: 10.1021/ja807430h.
A general strategy for the site-specific dual-labeling of proteins for single-molecule fluorescence resonance energy transfer is presented. A genetically encoded unnatural ketone amino acid was labeled with a hydroxylamine-containing fluorophore with high yield (>95%) and specificity. This methodology was used to construct dual-labeled T4 lysozyme variants, allowing the study of T4 lysozyme folding at single-molecule resolution. The presented strategy is anticipated to expand the scope of single-molecule protein structure and function studies.
本文提出了一种用于单分子荧光共振能量转移的蛋白质位点特异性双标记的通用策略。一种基因编码的非天然酮氨基酸被高产率（>95%）且特异性地用含羟胺的荧光团标记。该方法用于构建双标记的T4溶菌酶变体，从而能够在单分子分辨率下研究T4溶菌酶的折叠。预计所提出的策略将扩大单分子蛋白质结构和功能研究的范围。
