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Continuous Immobilized Recombinant Protein Production from E. coli Capable of Selective Protein Excretion: A Feasibility Study
Author(s) -
Georgiou G.,
Chalmers J. J.,
Shuler M. L.,
Wilson D. B.
Publication year - 1985
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1002/btpr.5420010114
Subject(s) - recombinant dna , plasmid , escherichia coli , bioprocess , bioreactor , biology , signal peptide , protein biosynthesis , gene , chemistry , biochemistry , paleontology , botany
Currently genetically engineered bacteria are economically attractive only for the production of high‐value, low‐volume‐proteins. For this technology to be useful for the manufacture of high volume proteins new bioprocessing concepts are necessary. A potential approach is to construct a plasmid containing an inducible promoter and a hybrid gene containing a signal sequence fused to the gene for the protein to be produced. The plasmid is transformed into Escherichia coli. The plasmid‐host interaction is such that the host becomes “leaky” upon introduction. Since the signal sequence allows the recombinant protein to pass the cytoplasmic membrane, a “leaky cell” can allow selective excretion of the desired protein. If the “leaky” cell is immobilized and maintained in a resting state, the sustained continuous production of the target protein is possible. To test this concept we have used plasmid pKK in E. coli RB791 with β‐lactamase as the target protein. Production levels of at least 390 units β‐lactamase/L‐h can be sustained for at least 120 days. Peak productivities of 10,000 unitslL‐h have been obtained. The β‐lactamase constitutes approximately 40% of the protein in solution. Such systems could reduce recombinant protein production costs due both to the continuous nature of the bioreactor and the improved ease of protein recovery.