Lee Hyun Soo, Spraggon Glen, Schultz Peter G, Wang Feng
Department of Chemistry, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
J Am Chem Soc. 2009 Feb 25;131(7):2481-3. doi: 10.1021/ja808340b.
A metal-ion chelating amino acid, (8-hydroxyquinolin-3-yl)alanine, was genetically encoded in E. coli by an amber nonsense codon and corresponding orthogonal tRNA/aminoacyl-tRNA synthetase pair. The amino acid was incorporated into TM0665 protein, and the mutant protein was cocrystallized with Zn(2+) to determine the structure by SAD phasing. The structure showed a high occupancy of the heavy metal bound to the HQ-Ala residue, and the heavy metal provided excellent phasing power to determine the structure. This method also facilitates the de novo design of metalloproteins with novel structures and functions, including fluorescent sensors.
一种金属离子螯合氨基酸，（8-羟基喹啉-3-基）丙氨酸，通过琥珀色无义密码子以及相应的正交tRNA/氨酰tRNA合成酶对在大肠杆菌中进行遗传编码。该氨基酸被整合到TM0665蛋白中，并且突变蛋白与Zn(2+)共结晶以通过单波长反常散射（SAD）相位法确定结构。该结构显示重金属与HQ-Ala残基结合的占有率很高，并且重金属为确定结构提供了出色的相位能力。这种方法还促进了具有新颖结构和功能的金属蛋白的从头设计，包括荧光传感器。
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