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

A simple procedure for preparation of a cell-free protein synthesis system (23000 × g supernatant) from protoplast-like ‘slime’ mutant of Neurospora crassa is described. A variety of messenger RNAs of viral and cellular origin could be efficiently and faithfully translated in this system into proteins with M, as large as 180000. The importance of the 7-methylguanosine 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 potassium ions, mRNA and 23000 × g supernatant protein.