FOLDING OF AN ENZYME INTO AN ACTIVE CONFORMATION WHILE BOUND AS PEPTIDYL-TRANSFER-RNA TO THE RIBOSOME

Rhodanese bound to bacterial ribosomes as peptidyl-tRNA can be folded into an enzymatically active conformation by generating C-terminal ext ensions of the wild-type enzyme. Rhodanese was synthesized by coupled transcription/translation in a cell-free Escherichia coli system from plasmids containing the coding sequences for the wild-type enzyme or i ts C-terminally extended mutants. Two proteins with extensions of 23 a mino acids or longer were enzymatically active while bound to the ribo somes whereas wild-type protein and a 13-amino acid extension were not . All forms of the enzyme were active after termination and release of the full-length protein from the ribosomes. All five of the bacterial chaperones were required to substantially increase the specific enzym atic activity of the extended rhodanese while the nascent protein was bound to ribosomes. The results provide direct support for the hypothe sis that proteins acquire tertiary structure as they are formed in rib osomes.