RIBOSOMAL-BINDING SITE OF RELEASE FACTORS RF1 AND RF2 - A NEW TRANSLATIONAL TERMINATION ASSAY IN-VITRO

We have established a new in vitro assay for translational termination . It consists of 70 S ribosomes bound to a synthetic RNA minimessenger via interaction with P-site binding fMet-tRNA(f)(Met). If the A-site codon is a stop signal, release activity can be measured by quantifyin g hydrolyzed formylmethionine, Characteristics of this assay in terms of reaction time, ion concentration, release factor RF1 and RF2 concen tration, and competition with A-site-decoding tRNA are discussed. The new assay shows that polypeptide chain release activity is directly de pendent on the presence of a stop codon in the ribosomal A-site.