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The site-specific incorporation of p-iodo-L-phenylalanine into proteins for structure determination


A recently developed method makes it possible to genetically encode unnatural amino acids with diverse physical, chemical or biological properties in Escherichia coli1 and yeast2. We now show that this technology can be used to efficiently and site-specifically incorporate p-iodo-L-phenylalanine (iodoPhe) into proteins in response to an amber TAG codon. The selective introduction of the anomalously scattering iodine atom into proteins should facilitate single-wavelength anomalous dispersion3,4 experiments on in-house X-ray sources. To illustrate this, we generated a Phe153 → iodoPhe mutant of bacteriophage T4 lysozyme and determined its crystal structure using considerably less data than are needed for the equivalent experiment with cysteine and methionine. The iodoPhe residue, although present in the hydrophobic core of the protein, did not perturb the protein structure in any meaningful way. The ability to selectively introduce this and other heavy atom–containing amino acids into proteins should facilitate the structural study of proteins.

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Figure 1: Evolution of a mutant M. jannaschii TyrRS that specifically incorporates iodoPhe into proteins in response to an amber stop codon in E. coli.
Figure 2: Patterson maps and difference Fourier maps for the anomalous data sets.
Figure 3: Structures and representative electron density of the iodoPhe.

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We would like to thank Heidi Erlandsen and Raymond Stevens for technical assistance. J.X. thanks Jeremy Mills, Jun Yin and Yan Zhang for helpful discussions. This work is supported by a grant from the National Institutes of Health (GM62159) and the Skaggs Institute for Chemical Biology. This is manuscript number 16547-CH of the Scripps Research Institute.

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Correspondence to Glen Spraggon or Peter G Schultz.

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Xie, J., Wang, L., Wu, N. et al. The site-specific incorporation of p-iodo-L-phenylalanine into proteins for structure determination. Nat Biotechnol 22, 1297–1301 (2004).

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