Production of authentic human proapolipoprotein A-I in Escherichia coli: Strategies for the removal of the amino-terminal methionine
| Autor: | Alex Bollen, Kees Roobol, Pascal Gilles, J.-P. Guillaume, Francesca Varsalona, Nicole Moguilevsky |
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| Rok vydání: | 1993 |
| Předmět: |
Enteropeptidase
Signal peptide Recombinant Fusion Proteins Molecular Sequence Data DNA Recombinant Bioengineering Biology Protein Engineering medicine.disease_cause Applied Microbiology and Biotechnology Aminopeptidase Methionine Escherichia coli medicine Humans Amino Acid Sequence Protein Precursors Apolipoproteins A Apolipoprotein A-I Base Sequence General Medicine Periplasmic space biology.organism_classification Molecular biology Enterobacteriaceae Fusion protein Enzymes Biochemistry Evaluation Studies as Topic Bacterial outer membrane Biotechnology |
| Zdroj: | Journal of Biotechnology. 27:159-172 |
| ISSN: | 0168-1656 |
| DOI: | 10.1016/0168-1656(93)90105-v |
| Popis: | Several methods were compared with respect to the production of authentic, N-terminal methionine-free proapolipoprotein A-I in engineered Escherichia coli bacteria. A first approach consisted of treating the purified methionylated recombinant protein with an amino-peptidase, purified from Aeromonas proteolytica. A second series of strategies was based on the construction of proapo A-I encoding cassettes carrying built-in recognition sites suitable for specific in vitro cleavage of the products with kallikrein and enterokinase, respectively. Along the same line, a fusion between ubiquitin and proapo A-I was produced in E. coli with the prospect to achieve post-purification cleavage with yeast ubiquitin hydrolase. Finally, proapo A-I was fused to the signal peptide of the bacterial outer membrane protein, OmpA, aiming at an in situ conversion to authentic proapo A-I during secretion to the bacterial periplasm. The data showed that, out of these five systems, the OmpA signal peptide system and, to a lesser extent, the one involving the fusion to ubiquitin were the most efficient in yielding authentic proapo A-I from engineered Escherichia coli. |
| Databáze: | OpenAIRE |
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