Enhanced D-amino acid incorporation into protein by modified ribosomes

Journal of the American Chemical Society

Volumn 125, Issue 22, 2003, Pages 6616-6617

  Dedkova, Larisa M ^a   Fahmi, Nour Eddine ^a   Golovine, Serguei Y ^a   Hecht, Sidney M ^a  

^a UNIVERSITY OF VIRGINIA   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

DEXTRO AMINO ACID; PEPTIDYLTRANSFERASE; PROTEIN; TRANSFER RNA;

ARTICLE; CHEMICAL MODIFICATION; CODON; GENE MUTATION; NONHUMAN; PROTEIN EXPRESSION; PROTEIN PROTEIN INTERACTION; PROTEIN SYNTHESIS; RIBOSOME; RNA SEQUENCE; SEQUENCE ANALYSIS;

AMINO ACIDS; CELL-FREE SYSTEM; ESCHERICHIA COLI; LUCIFERASES; MUTATION; PROTEIN BIOSYNTHESIS; PROTEINS; RIBOSOMES; RNA, MESSENGER; RNA, RIBOSOMAL, 23S; RNA, TRANSFER, AMINO ACYL; RNA, TRANSFER, MET; TETRAHYDROFOLATE DEHYDROGENASE;

EID: 0038547955     PISSN: 00027863     EISSN: None     Source Type: Journal    
DOI: 10.1021/ja035141q     Document Type: Article

References (24)

1. Hecht, S. M. Acc. Chem. Res. 1992, 25, 545.
2. Noren, C. J.; Anthony-Cahill, S. J.; Griffith, M. C.; Schultz, P. G. Science 1989, 244, 182.
3. Eisenhauer, B. M.; Hecht, S. M. Biochemistry 2002, 41, 11472 and references therein.
4. (a) Kreil, G. Annu. Rev. Biochem. 1997, 66, 337.
5. (b) Friedman, M. J. Agric. Food. Chem. 1999, 47, 3457.
6. Soyez, D.; Toullec, J.-Y.; Ollivaux, C.; Géraud, C. J. Biol. Chem. 2000, 275, 37870.
7. Luo, L.; Kohli, R. M.; Onishi, M.; Linne, U.; Marahiel, M. A.; Walsh, C. T. Biochemistry 2002, 41, 9184.
8. (a) Heckler, T. D.; Roesser, J. R.; Xu, C.; Chang, P. I.; Hecht, S. M. Biochemistry 1988, 27, 7254.
9. (b) Roesser, J. R.; Xu, C.; Payne, R. C.; Surratt, C. K.; Hecht, S. M. Biochemistry 1989, 28, 5185.
10. (c) Bain, J. D.; Diala, E. S.; Glabe, C. G.; Wacker, D. A.; Lyttle, M. H.; Dix, T. A.; Chamberlin, A. R. Biochemistry 1991, 30, 5411.
11. (a) Thompson, J.; Kim, D. F.; O'Connor, M.; Lieberman, K. R.; Bayfield, M. A.; Gregory, S. T.; Green, R.; Noller, H. F.; Dahlberg, A. E. Proc. Natl. Acad. Sci. U.S.A. 2002, 98, 9002.
12. (b) O'Connor, M.; Lee, W.-C.; Mankad, A.; Squires, C. L.; Dahlberg, A. E. Nucleic Acids Res. 2001, 29, 710.
13. Sawano, A.; Miyawaki, A. Nucleic Acids Res. 2000, 28, e78. Plasmid pUCrrnB, used in the mutagenesis procedure, was constructed by incorporation of the rrnB operon, excised from plasmid pNot by Kpnl and BamHI restriction endonucleases, and introduced into high-copy vector pUC18 under the control of an IPTG-induced Lac promoter.
14. O'Connor, M.; Dahlberg, A. E. J. Mol. Biol. 1995, 254, 838. The latter region plays an important role in translation.
15. 8a
16. 10 Three of six mutants had the same sequence, CUGGAG, instead of UGAUAC (wild-type). While the 23S rRNA sequences in Table 1 were inferred from DNA sequencing, the presence of the UUGUA sequence at positions 2447-2451 of the first mutant in the table was verified by hybridization of a radiolabeled DNA probe to the isolated 23S rRNA.
17. CUA. The initial rrnB gene used in this work contained one point mutation in the 23S rRNA gene (A1061T), which confers thiostrepton resistance (Spahn, C. M. T.; Remme, J.; Schafer, M. A.; Nierhaus, K. H. J. Biol. Chem. 1996, 271, 32849). Therefore, the levels of plasmid-born ribosome in all S-30 preparations were estimated from the level of protein synthesis in the presence of this antibiotic and verified by the primer extension method (Table 1); they varied from 36 to 63%. The translational fidelity of all S-30 preparations was estimated by the relative enzyme activity of E. coli DHFR synthesized in the presence of thiostrepton, which was ∼80-85% of DHFR prepared using wild-type ribosomes.
18. CUA. The initial rrnB gene used in this work contained one point mutation in the 23S rRNA gene (A1061T), which confers thiostrepton resistance (Spahn, C. M. T.; Remme, J.; Schafer, M. A.; Nierhaus, K. H. J. Biol. Chem. 1996, 271, 32849). Therefore, the levels of plasmid-born ribosome in all S-30 preparations were estimated from the level of protein synthesis in the presence of this antibiotic and verified by the primer extension method (Table 1); they varied from 36 to 63%. The translational fidelity of all S-30 preparations was estimated by the relative enzyme activity of E. coli DHFR synthesized in the presence of thiostrepton, which was ∼80-85% of DHFR prepared using wild-type ribosomes.
19. Lodder, M.; Golovine, S.; Laikhter, A. L.; Karginov, V. A.; Hecht, S. M. J. Org. Chem. 1998, 63, 794. D-Methionine and D-phenylalanine were treated with L-amino acid oxidase prior to attachment to the suppressor tRNA to destroy any residual L-amino acid.
20. CUA.
21. CUA's in a protein synthesizing system programmed with DHFR mRNA containing a UAG codon at position 42. The derived proteins were subjected to CNBr mapping, which verified the incorporation of D-methionine.
22. (a) Bystroff, C.; Oatley, S. J.; Kraut, J. Biochemistry 1990, 29, 3263.
23. (b) Murphy, D. J.; Benkovic, S. J. Biochemistry 1989, 28, 3025.
24. Branchini, B. R.; Magyar, R. A.; Murtiashaw, M. H.; Anderson, S. M.; Zimmer, M. Biochemistry 1998, 37, 15311.