Post-translational regulation of protein activity with small-molecule-controlled peptide splicing

ORGN 390

Georgios Skretas, gskretas@princeton.edu, Department of Chemical Engineering, Princeton University, Engineering Quadrangle, Princeton University, Princeton, NJ 08544 and David W. Wood, dwood@princeton.edu, Departments of Chemical Engineering and Molecular Biology, Princeton University, Engineering Quadrangle, A213 Olden Street, Princeton, NJ 08544.
Inteins are protein splicing elements that excise themselves post-translationally from a variety of protein hosts. Intein insertion abolishes, in general, the activity of its host protein, which is subsequently restored upon intein excision. Thus, inteins could be used as molecular “switches” for the control of arbitrary target proteins. Based on rational design, a chimeric intein has been constructed whose splicing activity is conditionally triggered in vivo by the presence of thyroid hormone. This chimera was used to demonstrate that different proteins can be inactivated by intein insertion and then reactivated by the addition of thyroid hormone or synthetic analogues in a dose-dependent manner. A combination of rational protein engineering and directed evolution was then employed to evolve inteins whose splicing activity is inhibited by the presence of synthetic estrogen ligands. Finally, the possibility of using small-molecule-controlled intein splicing to regulate protein function in higher eukaryotes will be discussed.
 

Proteins, Peptides, Amino Acids, and Enzymes
8:00 AM-11:40 AM, Tuesday, 15 March 2005 Convention Center -- Room 9, Oral

Division of Organic Chemistry

The 229th ACS National Meeting, in San Diego, CA, March 13-17, 2005