SummaryThe Arabidopsis thaliana lysyl tRNA synthetase (AtKRS) structurally and functionally resembles the wellcharacterized prokaryotic class IIb KRS, including the propensity to aminoacylate tRNA Lys with suboptimal identity elements, as well as non-cognate tRNAs. Transient expression of AtKRS in carrot cells promotes aminoacylation of such tRNAs in vivo and translational recoding of lysine at nonsense codons. Stable expression of AtKRS in Zea mays causes translational recoding of lysine into zeins, significantly enriching the lysine content of grain.

A new photo-cross-linking dU analog, 5-[4-(3-(trifluoromethyl)-3H-diazirin-3-yl)phenyl]-2'-deoxyuridine, was synthesized and incorporated into the recognition site of EcoRII and MvuI restriction-modification enzymes. The resulting base-modified 14-mer substrate was tested for cross-linking to these enzymes. Cross-linking is effected by irradiation of the enzyme-substrate complexes at 366 nm.

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Mutation of the Arabidopsis thaliana tRNA (Trp)(CCA) anticodon or of the A73 discriminator base greatly diminishes in vitro aminoacylation with tryptophan, indicating the importance of these nucleotides for recognition by the plant tryptophanyl-tRNA synthetase. Mutation of the tRNA (Trp)(CCA) anticodon to CUA so as to translate amber nonsense codons permits tRNA (Trp)(CCA) to be aminoacylated by A.thaliana lysyl-tRNA synthetase. Thus, translational suppression by tRNA (TRP)(CCA) observed in plant cells includes significant incorporation of lysine into protein.