+44-7360-538437- Biomedical Research (2011) Volume 22, Issue 3
Active recombinant Reverse Transcriptase Domain of human Hepatitis B Virus Polymerase
Hepatitis B virus polymerase plays a critical role during HBV life cycle, and polymerase/reverse transcriptase (RT) activities are critical for HBV-pol during viral replication. To investigate RT do-main of human HBV polymerase, a 5’ end Polyhistidine tagged RT DNA (304-693 amino acids) of HBV-pol was successfully expressed in Escherichia coli. Recombinant RT was purified in native condition employing Ni-NTA affinity column. Purified RT showed a stable reverse transcriptase ac-tivity and a much stronger DNA polymerase activity, compared to RT expressed in rabbit reticulo-cyte lysate coupled transcriptase-translation system. We present a new simplified way of obtaining active RT protein using the Escherichia coli expression and Reticulocyte lysate system. The purified RT was a stable protein and showed a low selective polymerase activity. Computer modeling results also indicated that RT domain banded to nucleotide substrate in a loose mode.
Author(s): Dipendra Raj Pandeya, Yu Yang, Seong-Tshool Hong