57 – Photoaffinity Labeling of Ribosomes

Publisher Summary The chapter discusses the photoaffinity of ribosomes. Affinity-labeling studies on the ribosome have already provided evidence as to the locations of the peptidyltransferase center, the mRNA codon site, and the binding sites for several antibiotics and protein cofactors involved in protein synthesis. The affinitylabeling approach, by which one forms a covalent bond between a ligand (usually radioactive) and its receptor site, permitting, on subsequent analysis, identification of the components of the ligand binding site, has yielded important information toward this end. The merits of this approach are that it can be applied to the intact ribosome, the data obtained are easily interpretable, and the sites of incorporation can in principle be determined to any desired level of detail, from individual proteins or extended RNA regions to individual amino acid or nucleotide residues. The chapter discusses covalent bond formation in affinity-labeling. The most common procedure is to prepare a photolabile or electrophilic derivative of the ligand whose binding site is being studied.