Akira Kaji
professor, University of Pennsylvania, USA
Title: The major action of Ribosome Recycling Factor (RRF) is to release mRNA from spent ribosomes—Use of this reaction for quick screening of specific antibiotic against RRF
Biography
Biography: Akira Kaji
Abstract
Protein synhesis has four steps, initiation, elongation of peptide chain, termination and the recycling of the spent ribosomes, mRNA and tRNA. The last step was discovered by our laboratories and catalyzed by a unique protein called ribosome recycling factor (RRF). The recycling step consists of three reactions, release of mRNA, tRNA and splitting of ribosomes. With the use of fluorescnt labeled mRNA similar to the natural mRNA and labeled tRNA, we demonstrate in vitro that the major action of RRF is to release of mRNA and not the splitting of ribosomes into subunits. This corrects the general misconcept that the major action of RRF is to split the ribosomes into subunits. The order of events with the naturally occuring substrate of RRF, the chain of events is release of tRNA, mRNA followed by the splitting of ribosomes. The release of mRNA is not dependent on the spliting of ribosomes. The in vitro results are supported by in vivo experiments where we used the tanlational coupling followed by the reporter gene expression (beta galactosidase expression). Using the basic reaction of RRF, release of ribosomes from mRNA, we developed a new screening system for the inhibitor of RRF. In this system, the inhibition of RRF reaction, makes ribosome stay on the mRNat the termination codon but start translating downstream which is linked to GFP. (222 words). We show that this screening method functions by the use of known specific inhibor of RRF, low concentration of fusidic acid. The assay method is simple and can be performed in 96 hole plate overnight. We look forward to find collaborators who has an access to the collection of possible inhibitors.