Selection of Modified Ribosomes Using Novel Puromycins

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Project Summary The long term goals of this research involve modification of the peptidyltransferase (PTC) center in bacterial ribosomes to enable the incorporation of very unusual amino acids not recognized by wild- type ribosomes. In recent years, we have described the selection of modified ribosomes and their use in incorporating D-amino acids, beta amino acids, dipeptides and dipeptidomimetric cassettes into proteins. During the four years of proposed research, the focus of efforts will be on the selection of ribosomes capable of introducing phosphorylated and glycosylated amino acids into specific proteins. The utility of this approach will be explored by analyzing the altered function of proteins into which phosphates and carbohydrates have been introduced, and by expressing a phosphorylated DNA binding protein in E. coli.