ENHANCED EXPRESSION OF THE YEAST URE2 PROTEIN IN ESCHERICHIA-COLI - THE EFFECT OF SYNONYMOUS CODON SUBSTITUTIONS AT A SELECTED PLACE IN THEGENE

The expression of the yeast Ure2 protein and its two N- and C-terminal HA-(YPYPVDYA) epitope and His-tag fusions has been enhanced in E. col i by selected silent mutagenesis of the URE2 gene. The two Arg-AGA cod ons at positions 253 and 254 of the URE2 gene coding sequence were exc hanged by CGT codons accordingly. This has allowed an increased yield (up to 100-fold) of the full-length protein synthesized. Western blott ing with HA-epitope-specific antibodies using N- and C-terminal Ure2p- HA(epitope)-His-tag fusion constructs confirmed the integrity of the r ecombinant proteins. The N-(C-) terminal tagged proteins were shown to possess biological activity of the natural Ure2 protein.