Genetic Incorporation of a Small, Environmentally Sensitive, Fluorescent Probe into Proteins in Saccharomyces cerevisiae

摘要

The introduction of fluorescent probes into proteins provides a powerful tool to study protein structure and function either in vitro or in vivo. One particularly useful method involves genetically fusing the protein of interest to fluorescent proteins. However, the size of naturally occurring fluorescent proteins (>20 kDa) and the typical requirement for C- or N-terminal fusions limit their utility as a local probe of structure and in some cases can perturb the structure or function of a protein. A variety of other methods have been developed to introduce fluorescent probes into protein in vitro and in vivo including the selective chemical or enzymatic modification of specific recognition motifs, the use of intein-based semisynthetic methods, and in vitro protein translation6 with chemically aminoacylated tRNAs.