Genetic incorporation of a small, environmentally sensitive, fluorescent probe into proteins in Saccharomyces cerevisiae.
Here, we report that the fluorescent amino acid, 3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid (Anap), can be genetically incorporated into proteins in yeast with excellent selectivity and efficiency by means of an orthogonal tRNA/aminoacyl-tRNA synthetase pair. This small, environmentally sensitive fluorophore was site-specifically incorporated into Escherichia coli glutamine binding protein and used to directly probe local structural changes caused by ligand binding. The small size of Anap and the ability to introduce it by simple mutagenesis at defined sites in the proteome make it a useful local probe of protein structure, molecular interactions, protein folding, and localization.
Lee
Hyun Soo
HS
Department of Chemistry, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Guo
Jiantao
J
Lemke
Edward A
EA
Dimla
Romerson D
RD
Schultz
Peter G
PG
D026921:Amino Acid Transport Systems, Neutral / Q000235:genetics; D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins / Q000235:genetics; D005456:Fluorescent Dyes / Q000737:chemistry*; D015202:Protein Engineering; D012343:RNA, Transfer / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics*; D015091:beta-Alanine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000235:genetics
C546053:3-(6-acetylnaphthalen-2-ylamino)-2-aminopropanoic acid; D026921:Amino Acid Transport Systems, Neutral; D029968:Escherichia coli Proteins; D005456:Fluorescent Dyes; D029701:Saccharomyces cerevisiae Proteins; C528486:glnH protein, E coli; D015091:beta-Alanine; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases
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