Direct expression of hepatitis B surface antigen gene in E. coli.

Area of Science:

• Molecular Biology
• Virology
• Biotechnology

Background:

• Hepatitis B virus (HBV) poses a significant global health challenge.
• Hepatitis B surface antigen (HBsAg) is a key viral protein and a target for vaccines and diagnostics.
• Understanding HBsAg gene structure-function relationships is crucial for biotechnological applications.

Purpose of the Study:

• To investigate the impact of truncating the HBsAg gene on its expression and function.
• To compare the expression levels of full-length and truncated HBsAg in a recombinant system.
• To analyze the polypeptide products of different HBsAg gene constructs.

Main Methods:

• Preparation of DNA fragments encoding full-length and truncated HBsAg genes from HBV/adw genome.
• Insertion of these fragments into an expression vector (pTRP801) to create recombinant plasmids (pTRP SS-6, pTRP SS-39, pTRP SS-50).
• Transformation of bacteria and analysis of recombinant growth, HBsAg expression, and minicell polypeptide formation.

Main Results:

• Recombinant bacteria with the full-length HBsAg gene (pTRP SS-6) showed inhibited growth and low HBsAg expression.
• Transformants with truncated HBsAg genes (pTRP SS-39, pTRP SS-50) exhibited no growth inhibition and considerable HBsAg expression.
• Minicells produced specific polypeptides of approximately 24 K, 23 K, and 22 K daltons for pTRP SS-6, pTRP SS-39, and pTRP SS-50, respectively.

Conclusions:

• Truncation of the NH2-terminal hydrophobic domain of the HBsAg gene significantly enhances its expression in recombinant systems.
• The removal of this domain appears to alleviate growth inhibition in host cells.
• These findings have implications for optimizing HBsAg production for vaccine and diagnostic development.