Abstract
The genes of 23 phage T5 amber suppressor tRNAs were constructed using site-directed mutagenesis and were cloned in plasmid pGFIB1 under control of the lipoprotein gene constitutive promoter. Suppression efficiency was measured in vivo after introducing recombinant plasmids into E. coli strains bearing mutations in the lacI gene. Strong amber suppressors (not less than 40% efficiency of the wild type) were obtained with tRNAs specific for amino acids Ala, Gln, Gly, His, Lys, Ser, and Tyr. Suppressor activity was considerably enhanced by additional substitutions which restored pairing in the D stem (tRN AAspCUA) and in the T stem (tRN AGluCUA) outside the anticodon, as well as by substitution U32C in the anticodon loop of tRN AArgCUA .
Original language | English (US) |
---|---|
Pages (from-to) | 846-853 |
Number of pages | 8 |
Journal | Molecular Biology |
Volume | 32 |
Issue number | 6 |
State | Published - Nov 1 1998 |
Keywords
- Amber suppressor tRNA
- Phage T5
- Site-directed mutagenesis
- Suppression efficiency
ASJC Scopus subject areas
- Biophysics
- Structural Biology