HIGH-FIDELITY TRANSLATION OF RECOMBINANT HUMAN HEMOGLOBIN IN ESCHERICHIA-COLI

Coexpression of di-alpha-globin and beta-globin in Escherichia coli in the presence of exogenous heme yielded high levels of soluble, functi onal recombinant human hemoglobin (rHb1.1), High-level expression of r Hb1.1 provides a good model for measuring mistranslation in heterologo us proteins. rHb1.1 does not contain isoleucine; therefore, any isoleu cine present could be attributed to mistranslation, most likely mistra nslation of one or more of the 200 codons that differ from an isoleuci ne codon by 1 bp, Sensitive amino acid analysis of highly purified rHb 1.1 typically revealed less than or equal to 0.2 mol of isoleucine per mol of hemoglobin. This corresponds to a translation error rate of le ss than or equal to 0.001, which is not different from typical transla tion error rates found for E. coli proteins. Two different expression systems that resulted in accumulation of globin proteins to levels equ ivalent to similar to 20% of the level of B. coli soluble proteins als o resulted in equivalent translational fidelity.