Rewriting the universal genetic code

ORGN 282

Virginia W. Cornish1, Zhongping Tan, zt22@columbia.edu1, Lucas Gartenmann Dickson, lg2239@columbia.edu1, Phillip Effraim, pre2001@columbia.edu1, Joshua Avins, jla2113@columbia.edu1, Stephen C. Blacklow, sblacklow@rics.bwh.harvard.edu2, and Anthony Forster, a.forster@vanderbilt.edu3. (1) Department of Chemistry, Columbia University, 3000 Broadway, Havemeyer Hall, MC 3111, New York, NY 10027, (2) Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, NRB 652E, 77 Avenue Louis Pasteur, Boston, MA 02115, (3) Department of Pharmacology and Vanderbilt Institute of Chemical Biology, Vanderbilt University, Preston Research Building, Room 459, Vanderbilt University Medical Center, 23rd Avenue South at Pierce, Nashville, TN 37232
The ribosome is unique among Nature's biosynthetic machines in that the catalytic center is distinct from the substrate recognition pocket. This separation of catalysis and substrate binding suggests the ribosome may be uniquely well suited to manipulation for the synthesis of novel polymers. Indeed in 1971, Rich and Fahnestock demonstrated that the protein biosynthetic machinery could synthesize polymers containing a random mixture of ester and amide linkages using misacylated hydroxy-Phe-tRNAPhe. With the development of an efficient method for the chemoenzymatic synthesis of aminoacyl-tRNAs, several groups then used misacylated suppressor tRNAs for site-directed mutagenesis with unnatural amino acids. Recently, we demonstrated that a sense codon reassignment strategy with a pure translation system allowed translation of multiple, adjacent sense codons with synthetic aminoacyl-tRNA substrates. This approach and related approaches breaking codon degeneracy open the possibility of using the protein biosynthetic machinery for template encoded synthesis of novel backbone polymers of defined length and composition with a pool of synthetic acyl-tRNAs. Here we present results further exploring the tolerance of the prokaryotic translational machinery to backbone analog substrates. We present biochemical studies aimed at understanding the mechanistic basis for the amino acid substrate specificity of the translational machinery. Building from these results, we then further exploit the translational machinery for the synthesis of oligomers other than polypeptides.