Studies on Translocation: III. CONDITIONS NECESSARY FOR THE FORMATION AND DETECTION OF A STABLE RIBOSOME-G FACTOR-GUANOSINE DIPHOSPHATE COMPLEX IN THE PRESENCE OF FUSIDIC ACID

Abstract

The conditions necessary for the formation and detection of a ribosome-G factor (the soluble translocation protein)-GDP complex in the presence of fusidic acid have been examined. Complex detection has been accomplished by sucrose density gradient sedimentation and both gel and Millipore filtration. The latter two methods reveal a good correlation, as a function of fusidic acid concentration, between the amount of isolatable complex and the inhibition of the GTPase activity of the G factor-ribosome system by the antibiotic. Complex formation requires Mg2+ (10 to 20 mm) but apparently does not require a monovalent cation. The binding of nucleotide requires and is limited by both ribosomes and G factor. An examination of the stoichiometry of complex formation was carried out by varying ribosomes and G factor. The results of these experiments, on the basis of certain assumptions, are interpreted to indicate the formation, in the presence of fusidic acid, of a relatively stable ternary complex involving ribosomes, G factor, and GDP in a molar ratio of 1:1:1.