The Cell‐Free Protein Synthesis System from the ‘Slime’ Mutant of Neurospora crassa

Abstract

A simple procedure for preparation of a cell-free protein synthesis system (23,000 .times. g supernatant) from the protoplast-like slime mutant of N. crassa is described. A variety of mRNA of viral and cellular origin could be efficiently and faithfully translated in this system into proteins with MW as large as 180,000. The importance of the 7-methylguanosine [m7G] cap for mRNA translation in the Neurospora system was studied in detail using the cap analogs and chemically decapped messengers. As in the case of reticulocyte lysate or wheat germ extract, the extent of m7G requirement for mRNA translation in a fungal extract strongly depended on translation conditions such as incubation temperature or concentration of K ions, mRNA and 23,000 .times. g supernatant protein.