Initiation of protein synthesis in yeast: binding of Met-tRNAi

Conditions for the binding of Met-tRNAi to 40 s ribosomal subunits and to proteins isolated out of the yeast ribosomal KCl wash were investigated. Sucrose density gradient experiments revealed that binding of Met-tRNAi to 40 s ribosomal subunits was catalyzed in a AUG and GTP dependent reaction. Binding of Met-tRNAi to proteins of the ribosomal KCl wash as assayed by the Millipore filter technique was found to be independent of AUG, GTP and 40 s ribosomal subunits. Additions of GTP yielded only slight stimulation, whereas Mg2+ caused dissociation of complexes. It was concluded that these reactions were most likely catalyzed by initiation factor eIF-2 although stimulation by GTP did not occur.