Abstract
Synthesis of bacterial proteins on the ribosome starts with a formylated methionine. Removal of the N-terminal formyl group is essential and is carried out by peptide deformylase (PDF). Deformylation occurs co-translationally, shortly after the nascent-chain emerges from the ribosomal exit tunnel, and is necessary to allow for further N-terminal processing. Here we describe the kinetic mechanism of deformylation by PDF of ribosome-bound nascent-chains and show that PDF binding to and dissociation from ribosomes is rapid, allowing for efficient scanning of formylated substrates in the cell. The rate-limiting step in the PDF mechanism is a conformational rearrangement of the nascent-chain that takes place after cleavage of the formyl group. Under conditions of ongoing translation, the nascent-chain is deformylated rapidly as soon as it becomes accessible to PDF. Following deformylation, the enzyme is slow in releasing the deformylated nascent-chain, thereby delaying further processing and potentially acting as an early chaperone that protects short nascent chains before they reach a length sufficient to recruit other protein biogenesis factors.