Purification of Plant Translation Factors for Reconstitution of Protein Synthesis

The extract from washed wheat embryos provides a powerful and versatile method for cell-free protein synthesis based on the accumulated DNA sequence information. Although the extract contains everything needed for protein synthesis, some of the required components may be uncharacterized, and some components of the extract do not directly involve in, or even inhibit protein synthesis. This has hampered us from detailed analyses of the translation reaction. Therefore, we have decided to make an attempt to construct a reconstituted protein synthesis system, which would be useful for better understanding of the mechanisms supporting eukaryotic protein synthesis and its regulation and would probably be applicable to synthetic biology and artificial manipulation of the translational apparatus. In the present study, we fractionated an extract from crude wheat germ according to published protocols to obtain the fractions containing the eukaryotic elongation factors (eEFs) 1A, 1B, and 2. eEF1A and eEF2 supported polyphenylalanine synthesis.