A fusion plasmid for the synthesis of lipopeptide-antigen chimeras in Escherichia coli
| Autor: | Hélène Bergeron, Maureen D. O'Connor-McCourt, Clement Rioux, Liping Lin, Peter C. K. Lau, Suzanne Grothe |
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| Rok vydání: | 1992 |
| Předmět: |
Signal peptide
Lipoproteins Recombinant Fusion Proteins Genetic Vectors Molecular Sequence Data Colicins Peptide Protein Sorting Signals Biology Origin of replication medicine.disease_cause Tyrosine Phosphorylation Site chemistry.chemical_compound Plasmid Bacterial Proteins Escherichia coli Genetics medicine Amino Acid Sequence Antigens Peptide sequence chemistry.chemical_classification Base Sequence Lipopeptide General Medicine Alkaline Phosphatase Molecular biology ErbB Receptors chemistry Plasmids |
| Zdroj: | Gene. 116:13-20 |
| ISSN: | 0378-1119 |
| DOI: | 10.1016/0378-1119(92)90623-w |
| Popis: | Lipopeptides are potential vaccine candidates with a built-in adjuvant property. To circumvent the present chemical route of synthesis for lipopeptide-antigen conjugates, the lipoprotein property of the pColE2-P9-encoded lysis protein, CelB, was used to create the bacterial fusion plasmid, pKLY3, to produce lipopeptide-antigen chimeras in Escherichia coli. Plasmid pKLY3 is a derivative of pKK233-2 with the origin of replication of the single-stranded DNA phage, fl. Under control of the promoter, ptrc, is the 5' end of the celB gene coding for a lipoprotein signal peptide and the first five amino acids (aa) (CQANY) of the mature lysis protein. As model systems for the synthesis of small and large lipopeptide-antigens, DNA sequences coding for the P2 peptide and E. coli alkaline phosphatase (PhoA) were fused in frame to the region of celB coding for a lipoprotein signal peptide and CQANY. P2 is a 12-aa peptide including a tyrosine phosphorylation site of the epidermal growth factor receptor (EGF-R). Inducible expression of stable lipohexapeptide CQANYV, lipo-CQANY-P2, and lipo-CQANYA-PhoA, was demonstrated. Similar expression was obtained for lipo-CIEGR-P2 and lipo-CIEGRA-PhoA in which IEGR is a cleavage recognition site for the blood coagulation factor, Xa. Like QANY, IEGR is predicted to form a beta-turn structure. The presence of a lipid moiety on the products was confirmed by demonstrating the incorporation of radioactive palmitic acid and inhibition of processing by globomycin. The lipid-modified peptides were also identified by incorporation of radioactive tyrosine, and the nature of the P2 peptide was verified immunologically.(ABSTRACT TRUNCATED AT 250 WORDS) |
| Databáze: | OpenAIRE |
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