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Expressing and purifying an anti-atherosclerosis polypeptide vaccine in Escherichia coli.
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- Author(s): Gaofu Q;Gaofu Q; Jie W; Xin Y; Roque RS; Jingjing L
- Source:
Protein expression and purification [Protein Expr Purif] 2004 Aug; Vol. 36 (2), pp. 198-206.
- Publication Type:
Journal Article; Research Support, Non-U.S. Gov't
- Language:
English
- Additional Information
- Source:
Publisher: Academic Press Country of Publication: United States NLM ID: 9101496 Publication Model: Print Cited Medium: Print ISSN: 1046-5928 (Print) Linking ISSN: 10465928 NLM ISO Abbreviation: Protein Expr Purif Subsets: MEDLINE
- Publication Information:
Publication: Orlando, FL : Academic Press
Original Publication: San Diego : Academic Press, c1990-
- Subject Terms:
- Abstract:
A chimeric polypeptide of TTP-CETPC was successfully expressed as inclusion bodies in Escherichia coli by fusing it with the C-terminus of asparaginase and a basic amino acid-rich peptide (KR). After partially purified by washing with 0.5% (v/v) Triton X-100 in 10 mM PB, the pellet was solubilized in 8 M urea. The solution was precipitated with single volume and double volumes of cold ethanol for removing impurities. The fusion protein in solution was precipitated with triple volumes of ethanol to increase purity and then hydrolyzed with 50 mM hydrochloric acid at 55 degrees C for 72 h. The TTP-CETPC polypeptide was released after the unique acid-labile aspartylprolyl bond in the fusion protein was cleaved by acid. After impurities were removed by adjusting the hydrolysis solution pH to 9.45 and then to 8.37, the TTP-CETPC polypeptide was further purified by DEAE-cellulose column. The TTP-CETPC containing fractions were eluted at 60-80 mM NaCl. The purified TTP-CETPC cysteines were oxidized to form into intermolecular disulfide bonded dimers for immunizing mice. Specific anti-CETP antibodies in mice serum were assayed by ELISA and Western blot to verify that antibodies against CETP had been successfully induced and lasted for more than seventeen weeks in vivo.
- Accession Number:
0 (Bacterial Proteins)
0 (CETP protein, human)
0 (Carrier Proteins)
0 (Cholesterol Ester Transfer Proteins)
0 (Glycoproteins)
0 (Recombinant Fusion Proteins)
0 (Tetanus Toxin)
0 (Vaccines, Subunit)
30KYC7MIAI (Aspartic Acid)
9DLQ4CIU6V (Proline)
QTT17582CB (Hydrochloric Acid)
- Publication Date:
Date Created: 20040714 Date Completed: 20050131 Latest Revision: 20131121
- Publication Date:
20250114
- Accession Number:
10.1016/j.pep.2004.04.005
- Accession Number:
15249041
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