Genetic Incorporation Of A Metal-ion Chelating Amino Acid Into Proteins As A Biophysical Probe

摘要

A simple, general method to selectively introduce metal ion binding sites into polypeptides would greatly facilitate the engineering of catalytic and redox active sites, radioisotope binding sites, structural elements, and spectroscopic probes into proteins. For example, recently it was shown that the genetic introduction of (2,2'-bipyridin-5-yl)alanine (Bpy-Ala) into the DNA binding protein catabolite activator protein generated a Cu~(2+) dependent oxidative DNA cleaving agent. Here we report that an amino acid derivative (HQ-Ala, 1) of the metal ion chelating group 8-hydroxyquinoline, which forms highly stable complexes with most transition metal ions and some lanthanides, can be genetically encoded in E. coli in response to the amber codon, TAG. Moreover, we show that addition of Zn~(2+) to HQ-Ala containing proteins results in the formation of both a fluorescent probe and of a heavy metal binding site for SAD phasing in protein crystallographic structure determination.