ABSTRACT
The effect of several oligodeoxynucleotides complementary to the fragments of yellow lupin tRNA(Phe) was tested in the aminoacylation of tRNA(Phe) and in the binding of Phe-tRNA(Phe) to poly-U-programmed eukaryotic ribosomes. Oligonucleotides tested in the aminoacylation test did not give any inhibition. Monomers and dimers did not have any significant influence on the binding assay, either. A different percentage of inhibition of the binding of Phe-tRNA to ribosomes has been observed for oligonucleotides. Heptamer complementary to the anticodon loop gave 100% inhibition of the binding reaction. However, the oligonucleotides complementary to both the anticodon loop and stem and longer than the heptamer were much less effective inhibitors. A high inhibitory effect was also observed for trimers and for the decamer complementary to the D-loop and CCA-end.
Subject(s)
Oligodeoxyribonucleotides/pharmacology , RNA, Transfer, Amino Acid-Specific/metabolism , RNA, Transfer, Phe/metabolism , Ribosomes/metabolism , Base Sequence , Molecular Sequence Data , Nucleic Acid Conformation , Oligodeoxyribonucleotides/chemical synthesis , Poly U/metabolism , RNA, Transfer, Phe/genetics , Seeds/metabolism , Structure-Activity RelationshipABSTRACT
Elongation factors 1 (EF-1) have been isolated from different plants: wheat, yellow lupine, blue lupine, Chinese cabbage and Norway maple. Antibodies for EF-1 from yellow lupine have been obtained in rabbits; antibodies for wheat EF-1 were elicited in mice. The immunological properties of EF-1 were assayed by the following methods: western blotting, double immunodiffusion and rocket immunoelectrophoresis. Our results suggest that one antigenic site is similar for all plant elongation binding factors tested. This epitope probably overlaps the centre of biological activity of EF-1, as was shown for wheat EF-1. The hypothesis concerning the potential presence of plant EF-1 as a subunit of turnip yellow mosaic virus RNA replicase (similar to prokaryotic EF-Tu in the Q beta RNA replicase system) has also been tested using immunotechniques as well as tests of biological activity, but has not been confirmed.
Subject(s)
Peptide Elongation Factors/immunology , Cross Reactions , Immunodiffusion , Molecular Weight , Mosaic Viruses/enzymology , Plants , RNA-Dependent RNA Polymerase/immunology , Species SpecificityABSTRACT
Three forms of elongation factor 1 (EF1A, EF1B and EF1C) were isolated from wheat germ; the preparation obtained by successive chromatography on DEAE-Sephadex A-50, hydroxylapatite and Sephadex G-200 showed about 90% purity. The molecular masses of these forms were about: 61 000, 48 000 and 12 500, respectively. EF1A was the only form which formed the ternary complex GTP-EF1-AA-tRNA and was the most active in binding of Phe-tRNA to the poly-U programmed ribosomes. Based on the data of amino acid analysis, N-terminal amino acid determination and previously described proteolysis (Pulikowska et al., Biochem. Biophys. Res. Commun., 1979, 91, 1011-1017), it is assumed that EF1C is a basic structural subunit of elongation factor EF1 from wheat germ, but only EF1A shows the conformation-dependent full biological activity.