Selective inactivation of amino acid acceptor and ribosome-binding activities of Escherichia coli tRNA by modification with cyanogen bromide

Abstract

1. Modification of Escherichia coli tRNA with cyanogen bromide caused a selective loss of acceptor activities for glutamic acid, lysine and glutamine. However, most tRNA's known to contain 4-thiouridine, which is susceptible to cyanogen bromide, retained their amino acid acceptor activities after the modification, indicating that modification of the 4-thiouridylate residue has no effect on amino acid acceptor activity. 2. Inactivation of glutamic acid acceptor activity was due to a modification of 5-methylaminomethyl-2-thiouridine located in the first position of the anticodon of tRNA Glu, showing that the anticodon region is related to the recognition site of aminoacyl-tRNA synthetase in this particular tRNA. 3. The binding abilities of [ 14C]tyrosyl-, [ 14C]histidyl- and [ 14C]lysyl-tRNA's to ribosomes were specifically inactivated by the modification, whereas those of [ 14C]arginyl-, [ 14C]isoleucyl-, [ 14C]glycyl-, [ 14C]phenylalanyl- and [ 14C]valyl-tRNA were not affected. It is likely that the inactivation of the ribosome-binding ability of tRNA is due to the modification of other minor nucleosides which are susceptible to cyanogen bromide rather than to modification of 4-thiouridine. 4. [ 14C]Tyrosyl-tRNA modified with CNBr formed a complex with poly (U 4,G) in the absence of ribosomes. This suggested that modification with cyanogen bromide caused a distortion of the conformational structure of tRNA Tyr.