title: Glycine to glutamic acid misincorporation observed in a recombinant protein expressed by Escherichia coli cells.
abstract: A novel amino acid misincorporation, in which the intended glycine (Gly) residues were replaced by a glutamic acid (Glu), was observed in a recombinant protein expressed by Escherichia coli. The misincorporation was identified by peptide mapping and liquid chromatography-tandem mass spectrometric analysis on proteolyzed peptides of the protein and verified using the corresponding synthetic peptides containing the misincorporated residues. Analysis of the distribution of the misincorporated residues and their codon usage shows strong correlation between this misincorporation and the use of rarely used codon within the E. coli expression system. Results in this study suggest that the usage of the rare codon GGA has resulted in a Glu for Gly misincorporation.
authors.affiliation: Department of Biologics Product & Process Development, Bristol-Myers Squibb Technical Operations, Princeton, New Jersey 08543, USA. yunping.huang@bms.com
authors.forename: Yunping
authors.initials: Y
authors.lastname: Huang
authors.forename: Brian
authors.initials: B
authors.lastname: O'Mara
authors.forename: Matthew
authors.initials: M
authors.lastname: Conover
authors.forename: Richard
authors.initials: R
authors.lastname: Ludwig
authors.forename: Jinmei
authors.initials: J
authors.lastname: Fu
authors.forename: Li
authors.initials: L
authors.lastname: Tao
authors.forename: Zheng Jian
authors.initials: ZJ
authors.lastname: Li
authors.forename: Siegfried
authors.initials: S
authors.lastname: Rieble
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Grace
authors.forename: Reb J
authors.initials: RJ
authors.lastname: Russell
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D003062:Codon; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D018698:Glutamic Acid / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D005998:Glycine / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D003062:Codon; D010446:Peptide Fragments; D011994:Recombinant Proteins; D018698:Glutamic Acid; D005998:Glycine