Heterologous expression, characterization and structural studies of a hydrophobic peptide from the HIV-1 p24 protein

• Published in: Peptides (New York, N.Y. : 1980) N.Y. : 1980), 2005-02, Vol.26 (2), p.243-249
• Main Authors: Castilho, Priscila V., Campana, Patricia T., Garcia, Assuero F., Beltramini, Leila M., Araújo, Ana Paula U.
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Circular Dichroism; Escherichia coli - genetics; Fluorescence spectroscopy; Fundamental and applied biological sciences. Psychology; Genetic Vectors; HIV Core Protein p24 - chemistry; Hydrophobic and Hydrophilic Interactions; Mass Spectrometry; p24 HIV-1; Peptides - chemistry; Peptides - genetics; Peptides - metabolism; Protein Structure, Secondary; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - metabolism; Recombinant peptide; Sequence Analysis, Protein; Solubility; Spectrometry, Fluorescence; Synthetic gene; Vertebrates: endocrinology
• ISSN: 0196-9781; 1873-5169
• DOI: 10.1016/j.peptides.2004.09.014

Proteins from the inner core of HIV-1, such as the capsid protein (p24), are involved in crucial processes during the virus life cycle. The p24 protein plays an active structural role in the Gag protein and in its mature form. This work describes the production of a peptide derived from the p24 C-terminal, TLRAEQASQEVKNWMTETLLVQNA, using recombinant technology. This region (p24-3) is involved in interfaces during the p24 dimerization, which occurs during capsid assembly. The p24-3 sequence was obtained by a synthetic gene strategy and inserted into the pET 32a expression vector to produce soluble fusion protein in Escherichia coli BL21(DE3). This strategy leads to an incorporation of three amino acid residues (AMA) in the N-terminal of the native sequence to form the recombinant p24-3 (rp24-3). The rp24-3 was purified by reverse phase chromatography to homogeneity, as inferred by mass spectrometry and protein sequence analysis. Structural studies using circular dichroism and steady-state fluorescence showed that the rp24-3 is structured by helical and beta elements. As a function of its hydrophobic character it can self-associate forming oligomers. We present in this paper the first development of a suitable expression system for rp24-3, which provides high amounts of the peptide. This strategy will allow the development of new antiviral (HIV) agents.