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Amino acid content of recombinant proteins influences the metabolic burden response
Author(s) -
Bonomo Jeanne,
Gill Ryan T.
Publication year - 2005
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.20436
Subject(s) - amino acid , escherichia coli , recombinant dna , gene , translation (biology) , biochemistry , biology , peptide sequence , protein biosynthesis , chemistry , microbiology and biotechnology , messenger rna
Recombinant protein production in Escherichia coli often results in a dramatic cellular stress response best characterized by a decrease in overall cell fitness. We determined that the primary sequence (the amino acid sequence) of the recombinant protein alone plays an important role in mitigating this response. To do so, we created two polypeptides, modeled after the 39‐40 amino acid Defensin class of proteins, which contained exclusively the five least (PepAA; His, Trp, Tyr, Phe, Met), or most (PepCO: Ala, Glu, Gln, Asp, Asn) abundant amino acids in E. coli . We determined that overexpression of PepAA resulted in a drastic decrease in growth rate compared to overexpression of PepCO, our model Defensin protein MGD‐1, or the 26 amino acid polypeptide contained within the pET‐3d vector backbone. We further determined, using Affymetrix E. coli gene chips, that differences among the whole‐genome transcriptional responses of these model systems were best characterized by altered expression of genes whose products are involved in translation, transport, or metabolic functions as opposed to stress response genes. Based on these results, we confirmed that translation efficiency was significantly reduced in cells overexpressing PepAA compared with the other model polypeptides evaluated. © 2005 Wiley Periodicals, Inc.