Optimized orthogonal translation of unnatural amino acids enables spontaneous protein double-labelling and FRET.

ABSTRACT: The ability to introduce different biophysical probes into defined positions in target proteins will provide powerful approaches for interrogating protein structure, function and dynamics. However, methods for site-specifically incorporating multiple distinct unnatural amino acids are hampered by their low efficiency. Here we provide a general solution to this challenge by developing an optimized orthogonal translation system that uses amber and evolved quadruplet-decoding transfer RNAs to encode numerous pairs of distinct unnatural amino acids into a single protein expressed in Escherichia coli with a substantial increase in efficiency over previous methods. We also provide a general strategy for labelling pairs of encoded unnatural amino acids with different probes via rapid and spontane

SUBMITTER: Wang K 

PROVIDER: S-EPMC4430801 | biostudies-literature | 2014 May

REPOSITORIES: biostudies-literature