Cell-free protein synthesis in lysates of Drosophila melanogaster cells

Abstract

A procedure is described for preparing cell-free proteins synthesizing lysates from D. melanogaster tissue culture cells and embryos. Preparation of translationally active lysates from tissue culture cells is dependent on the presence of rat liver supernatant during cell lysis to inhibit ribonuclease activity. After micrococcal nuclease treatment of the lysate, protein synthesis is dependent on the addition of exogenous mRNA. The fidelity of translation is very high. The conditions for optimal translation were determined. The effects on translation of a variety of supplements, including rat liver supernatant, were analyzed. The products of translation by the Drosophila lysate were compared with those of wheat germ extracts and of micrococcal nuclease treated rabbit reticulocyte lysates. Translation in vitro of bovine parathyroid hormone mRNA yielded 2 products tentatively identified as preproparathyroid hormone and proparathyroid hormone, and an unidentified 3rd product. Insect enzymes can apparently accurately process mammalian precursor proteins.