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The Skaggs Institute
for Chemical Biology


Scientific Report 2006




New Amino Acid Building Blocks


P.G. Schultz, E. Brustad, A. Galkin, J. Graziano, J. Grbic, D. Groff, W. Hur, J. Melnick, J. Mills, J. Xie

Almost all processes of living cells, from gene regulation and information processing to photosynthesis, are carried out by proteins. These large molecules are synthesized from 20 amino acid building blocks. Indeed, this set of 20 amino acids is the basis for the genetic code, the code that specifies the relationship between the nucleotide sequence of a gene and the amino acid sequence of a protein. This fact leads to the rather interesting question of why every form of life has the same set of building blocks. Why not 21 or more? Moreover, if we can add new amino acid building blocks to the genetic code, will we be able to create proteins or even organisms with enhanced chemical, physical, or biological properties?

We are addressing this issue by using a number of chemical and molecular biological methods to add new components to the protein biosynthetic machinery. Using this approach, we effectively expanded the genetic code of Escherichia coli and yeast by genetically encoding new amino acids (including photoaffinity labels, reactive amino acids, glycosylated amino acids, and amino acids with altered electronic and steric properties) in response to unique 3- and 4-base codons.

Currently, we are exploring additional amino acids with novel biological and physicochemical properties, other organisms, and a number of biomedical applications of this technology. In the past year, we have (1) genetically encoded a fluorescent amino acid, the infrared probe para-cyanophenylalanine, a variety of photoactivated amino acids, cysteine homologs, amino acids containing boronate or ferrocene side chains and sulfotyrosine; (2) expanded this method to mammalian cells by adapting orthogonal tRNA–aminoacyl tRNA synthetase pairs evolved in yeast; (3) optimized the expression of mutant proteins in E coli and yeast; and (4) solved structures of benzophenone- and bipyridyl-specific aminoacyl tRNA synthetases.

In the next year we will (1) continue our efforts to genetically encode amino acid analogs with altered backbones (α-hydroxy-, N-methyl-, and β-amino acids); (2) use a metal-binding acid to make a “T20” trimer to block HIV infection; (3) generate serum “carrier proteins” that can be modified with therapeutic peptides and short-lived small molecules; (4) generate libraries of peptides and proteins containing unnatural amino acids and screen the molecules for high-affinity ligands to various receptors; and (5) biosynthesize cyclic proteins and peptides by using amino acids with reactive side chains (phenylselenide, thioester).

In a separate project, we have been using genomics tools to identify genes that regulate key biological pathways and processes. One such process is the cell cycle, which controls the growth and division of cells throughout life. A detailed understanding of the global regulation of this fundamental process requires comprehensive identification of the genes and pathways involved in the various stages of the cycle. To this end, we have carried out a genome-wide analysis of the human cell cycle, cell size, and proliferation by using small interfering RNAs to target more than 95% of the protein-coding genes in the human genome. Analysis of more than 2 million images, acquired by quantitative fluorescence microscopy, indicated that depletion of 1152 genes strongly affected progression of the cell cycle. Functional studies of these genes in our laboratory and elsewhere will provide systems-level insight into both known and novel genes and to pathways that regulate the cell cycle. Some of the findings may provide new therapeutic approaches for the treatment of cancer.

Publications

Liu, C., Schultz, P.G. Recombinant expression of selectively sulfated proteins in Escherichia coli. Nat. Biotechnol. 24:1436, 2006

Liu, W., Brock, A., Chen, S., Chen, S., Schultz, P.G. The genetic incorporation of unnatural amino acids into proteins in mammalian cells. Nat. Methods, in press.

Mukherji, M., Bell, R., Supekova, L., Wang, Y., Orth, A.P., Batalov, S., Miraglia, L., Huesken, D., Lange, J., Martin, C., Sahasrabudhe, S., Reinhardt, M., Natt, F., Hall, J., Mickanin, C., Labow, M., Chanda, S.K., Cho, C.Y., Schultz, P.G. Genome-wide functional analysis of human cell-cycle regulators. Proc. Natl. Acad. Sci. U. S. A. 103:14819, 2006.

Schultz, K.C., Supekova, L., Ryu, Y., Xie, J., Perera, R., Schultz, P.G. A genetically encoded infrared probe. J. Am. Chem. Soc. 128:13984, 2006.

Wang, J., Xie, J., Schultz, P.G. A genetically encoded fluorescent amino acid. J. Am. Chem. Soc. 128:8738, 2006.

Xie, J., Schultz, P.G. A chemical toolkit for proteins: an expanded genetic code. Nat. Rev. Mol. Cell Biol. 7:775, 2006.

 

Peter Schultz, Ph.D.
Scripps Family Chair Professor

Schultz Web Site