Prokaryotic expression and purification of human HDAC6 and preliminary detection of its function

Objective To obtain active purified GST-HDAC6 protein by cloning histone deacetylase 6 (HDAC6). Methods The coding sequence of HDAC6 was amplified by PCR from human mammary cDNA library, and inserted into the prokaryotic expression vector pGEX-KG. The fusion protein was purified by GST-Sepharose 4B beads, and then identified by SDS-PAGE and Western blot analysis. The biological activity of purified GST-HDAC6 protein was detected by deacetylation experiment. Results From human breast cDNA library, we got a 3645-bp fragment and cloned it into pGEX - KG carrier successfully of which the sequence was in complete accord with target sequence. The target protein expressed by the sequence was with a molecular weight of 170 000(Mr) from e. coli DH5α. Then, we obtained the highly purified fusion protein GST - HDAC6 after purification. By deacetylation experimental verification, we found the protein showed great enzyme activity. Conclusion HDAC6 is successfully cloned and the active fusion protein is purified, which provide experimental basis for research in relationship between deacetylases and tumor.