Fig. 6.

Pentapeptide synthesis.

A. Experimental approach. Initiation complexes (1 μM) were mixed with four different purified aa-tRNAs (1.1 μM each) in the presence of EF-Tu (1.65 μM), EF-G (2 μM), GTP (1 mM), phosphoenolpyruvate (3 mM) and pyruvate kinase (0.1 μg μl⁻¹). In each cycle, only one aa-tRNA was ¹⁴C-labelled.

B. Binding of aa-tRNA (grey bars, monitored by nitrocellulose filtration) and formation of the pentapeptide (black bars, analysed by HPLC) on colicin E3-cleaved ribosomes, normalized to the respective values obtained with control ribosomes (white bars). About 80% of control ribosomes bound each aa-tRNA and formed the pentapeptide fMetValTyrIlePhe. Labels indicate the position in the peptide of the labelled amino acid being analysed in each row. nd, not determined.