title: High-fidelity translation of recombinant human hemoglobin in Escherichia coli.
abstract: Coexpression of di-alpha-globin and beta-globin in Escherichia coli in the presence of exogenous heme yielded high levels of soluble, functional recombinant human hemoglobin (rHb1.1). High-level expression of rHb1.1 provides a good model for measuring mistranslation in heterologous proteins. rHb1.1 does not contain isoleucine; therefore, any isoleucine present could be attributed to mistranslation, most likely mistranslation of one or more of the 200 codons that differ from an isoleucine codon by 1 bp. Sensitive amino acid analysis of highly purified rHb1.1 typically revealed < or = 0.2 mol of isoleucine per mol of hemoglobin. This corresponds to a translation error rate of < or = 0.001, which is not different from typical translation error rates found for E. coli proteins. Two different expression systems that resulted in accumulation of globin proteins to levels equivalent to approximately 20% of the level of E. coli soluble proteins also resulted in equivalent translational fidelity.
authors.affiliation: Somatogen, Inc., Boulder, Colorado 80301, USA. mweickert@ligand.com
authors.forename: M J
authors.initials: MJ
authors.lastname: Weickert
authors.forename: I
authors.initials: I
authors.lastname: Apostol
mesh_terms: D004926:Escherichia coli / Q000378:metabolism*; D006454:Hemoglobins / Q000032:analysis / Q000096:biosynthesis* / Q000302:isolation & purification; D006801:Humans; D007532:Isoleucine / Q000032:analysis; D014176:Protein Biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification; D014633:Valine / Q000031:analogs & derivatives / Q000032:analysis
chemical_list: D006454:Hemoglobins; D011994:Recombinant Proteins; D007532:Isoleucine; C005313:norvaline; D014633:Valine