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J Bacteriol. 1974 February; 117(2): 439-443
Copyright © 1974 American Society for Microbiology. All Rights Reserved.

Suppression of Glutamic Acid Codons by Mutant Glycine Transfer Ribonucleic Acid

^a Department of Biological Sciences, Stanford University, Stanford, California 94305

ABSTRACT

In previous mutational studies with mutant trpA46 (Gly [GGA] Glu [GAA] at position 211 of the tryptophan synthetase alpha chain) of Escherichia coli, no missense suppressors were detected. Such suppressors have now been obtained by single mutations in gly Vins, the structural gene for a GGA/G-reading, mutationally altered form of gly V transfer ribonucleic acid (tRNA) (tRNA^Gly which reads GGU/C). A trpA46 strain containing the gly Vins alteration was mutagenized with hydroxylamine, and suppressor mutations were detected in the prototrophs obtained. Eighteen independent suppressors were examined and shown to have alterations which map in the gly V region. Chromatography of the glycyl-tRNAs of one suppressed mutant on a benzoylated diethylaminoethyl-cellulose column revealed an alteration in the tRNA_ins^Gly peak. The trpA46 suppressor mutation thus appears to involve a change of tRNA_ins^Gly from a GGA/G (Gly) reader to a GAA (Glu) reader. Since this suppressor presumably retains the "wobble" pairing of gly Vins tRNA, it was used to select the conversion of GAU (Asp211) to GAG (Glu211) in the alpha chain. supD (serine-inserting amber suppressor) was then used to obtain the conversion of GAG (Glu211) to UAG211. Missense revertants of trpA (UAG211) are being isolated as a means of introducing new codons which can be used in the selection of additional missense suppressors.

¹ Present address: Section of Molecular Biology, The University of Texas M. D. Anderson Hospital and Tumor Institute, Texas Medical Center, Houston, Tex. 77025.