A GROWTH-DEFECTIVE KIRROMYCIN-RESISTANT EF-TU ESCHERICHIA-COLI MUTANTAND A SPONTANEOUSLY EVOLVED SUPPRESSION OF THE DEFECT

This study has investigated the cause of a. growth-defect phenotype of a mutation in the elongation factor EF-Tu from Escherichia coli. An M 13-based genetic retrieval system reported by Zeef and Bosch [Mol. Gen . Genet. 238 (1993) 252-260] was used to segregate and identify an ext remely growth-defective kirromycin-resistant (Kr-R) tufA mutation, enc oding Gln(124)-->Lys (Q124K), from a Kr-R parent strain. This original strain also contained mutations, 124com1 and 124com2, that appear to have evolved to suppress the Q124K tufA. mutation. In this communicati on we present these M13-based genetic experiments together with additi onal genetic and protein characterization experiments to clarify the b asis of this complementation. The data indicate that the serious growt h defect of Q124K originates from a defective GTP/GDP interaction. The GTP/GDP binding and GTP hydrolysis characteristics of EF-Tu Q124K wer e different from wild-type EF-Tu and especially of another Kr-R EF-Tu mutant A375T. In line with this, 124com1 specifically complemented EF- Tu Q124K, whereas the growth defects of strains containing EF-Tu mutat ed at aa 375 were aggravated. We also show that strains containing the segregated tufA Q124K mutation formed filaments.