Firefly luciferase was shown to be completely folded and thus enzymatically active immediately upon release from the ribosome [Kolb et al. (1994) EMBO J. 13, 3631–3637]. However, no luciferase activity was observed while full-length luciferase was attached to the ribosome as a peptidyl-tRNA, probably because the C-terminal portion of the enzyme is masked by the ribosome and/or ribosome-associated proteins. Here we have demonstrated that the ribosome-bound enzyme acquires the enzymatic activity when its C-terminus is extended by at least 26 additional amino acid residues. The results demonstrate that the acquisition of the final native conformation by a nascent protein does not need the release of the protein from the ribosome.
