Translation initiation by using various N-acylaminoacyl tRNAs.
Bioactive peptides isolated from natural sources have diverse acyl groups on the N-terminus. It is difficult to synthesize these peptides in vitro translation system because ribosomal peptide synthesis generally limits the N-terminal group to be N-formylmethionine (fMet). To overcome this restriction, we developed a novel methodology for the ribosomal synthesis of peptides having various terminal N-acyl groups with desired amino acids. In this methodology, two technologies, Flexizyme system consisting of artificial ribozymes and a reconstitute E. coli cell-free translation system (PURE system), were used. First, an amino acid carrying a desired N-acyl group was charged onto an initiation tRNA by the Flexizyme system. The addition of this N-acyl-aminoacyl-tRNA (N-acyl-aa-tRNA) to the PURE system allowed us to initiate the peptide synthesis with the designated N-acyl-amino acid. By means of this methodology, the translation was exclusively initiated by various N-terminal acyl groups as well as amino acids without contamination of N-formylmethionine.
Goto
Yuki
Y
Research Center of Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Tokyo, Japan.
Ashigai
Hiroshi
H
Sako
Yusuke
Y
Murakami
Hiroshi
H
Suga
Hiroaki
H
D049148:Aminoacylation; D002474:Cell-Free System; D004926:Escherichia coli / Q000235:genetics; D010442:Peptide Chain Initiation, Translational; D010455:Peptides / Q000737:chemistry*; D010649:Phenylalanine / Q000031:analogs & derivatives; D016337:RNA, Catalytic / Q000378:metabolism; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry* / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000737:chemistry / Q000378:metabolism*; D046249:Transfer RNA Aminoacylation
D010455:Peptides; D016337:RNA, Catalytic; D012346:RNA, Transfer, Amino Acyl; D012358:RNA, Transfer, Met; D010649:Phenylalanine