The polysomal mRNA from the cell-free system of the yeast Saccharomyces cerevisiae, in the absence of exogenous energy, binds to the 40S ribosomal subunit thus forming a 48S preinitiation complex which, with energy added, is converted into 80S initiation complex. By using ribosomes with a high affinity to polysomal mRNA (pmRNA) from an edeine-resistant mutant of S. cerevisiae in place of wild-type ribosomes, increased quantities of the 48S preinitiation complex are obtained. The pmRNA is found associated with several polypeptides having molecular masses of 115-98 kDa, 72 kDa, 60 kDa and 51 kDa. These polypeptides, labelled with 125I, interact with 40S and 80S ribosomes and are essential for the formation of the 48S and 80S initiation complexes inasmuch as deproteinized pmRNA alone cannot initiate the process. In addition, other polypeptides present in the cytosol are required to carry out the above-mentioned steps of protein synthesis.