%0 Journal
%T Construction of modified ribosomes for incorporation of D-amino acids into proteins
%A Larisa M Dedkova, Nour Eddine Fahmi, Serguei Y Golovine, Sidney M Hecht
%J Biochemistry
%V 45
%P 15541-15551
%D 2006
%R 10.1021/bi060986a
%X While numerous biologically active peptides contain D-amino acids, the elaboration of such species is not carried out by ribosomal synthesis. In fact, the bacterial ribosome discriminates strongly against the incorporation of D-amino acids from D-aminoacyl-tRNAs. To permit the incorporation of D-amino acids into proteins using in vitro protein-synthesizing systems, a strategy has been developed to prepare modified ribosomes containing alterations within the peptidyltransferase center and helix 89 of 23S rRNA. S-30 preparations derived from colonies shown to contain ribosomes with altered 23S rRNAs were found to exhibit enhanced tolerance for D-amino acids and to permit the elaboration of proteins containing D-amino acids at predetermined sites. Five specific amino acids in Escherichia coli dihydrofolate reductase and Photinus pyralis luciferase were replaced with D-phenylalanine and D-methionine, and the specific activities of the resulting enzymes were determined.