[Prokaryotic soluble expression, purification and function study of LEDGF/p75 protein]

Area of Science:

• Biochemistry
• Molecular Biology
• Virology

Background:

• HIV-1 integrase (IN) is essential for viral replication.
• The interaction between HIV-1 IN and LEDGF/p75 is a key target for anti-HIV drugs.

Purpose of the Study:

• To construct and express functional recombinant human lens epithelium-derived growth factor (LEDGF/p75) in E. coli.
• To establish a platform for screening inhibitors of the HIV-1 IN-LEDGF/p75 protein-protein interaction.

Main Methods:

• Synthesized and optimized the LEDGF/p75 gene for E. coli expression.
• Cloned the gene into a pGEX-4T-1 vector and transformed into E. coli BL21 (DE3).
• Purified the recombinant protein using Ni2+ affinity chromatography and analyzed via SDS-PAGE and ELISA.

Main Results:

• Successfully expressed high yields of soluble and stable recombinant LEDGF/p75 in E. coli.
• Confirmed the recombinant protein's ability to enhance HIV-1 IN strand transfer activity in vitro.
• Validated the protein's function using ELISA assays.

Conclusions:

• The developed recombinant LEDGF/p75 is suitable for functional assays.
• This work facilitates the development of a screening platform for novel anti-HIV therapeutics targeting the IN-LEDGF/p75 interaction.