Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
17816 | Enzyme and Microbial Technology | 2008 | 7 Pages |
Abstract
In this research, a recombinant high mobility group box 1 AB peptide with a 6-histidine tag (rHMGB1AB-His6) was produced and characterized as a carrier of nucleic acids. The human HMGB1AB cDNA was cloned by RT-PCR. The rHMGB1AB-His6 expression vector, pET21a-rHMGB1AB-His6, was constructed with the cDNA. In pET21a-rHMGB1AB-His6, the acidic box of the C-terminus of wild type HMGB1 was eliminated, since it had negative charges and interfered with the DNA and peptide interaction. pET21a-rHMGB1AB-His6 was transformed into the BL21 strain. rHMGB1AB-His6 was produced by IPTG induction and purified using a nickel column. A gel retardation assay showed that plasmid DNA (pDNA) was completely retarded at a 1:1 weight ratio. The complex size was approximately 150Â nm at a 1:10 weight ratio (pDNA/rHMGB1AB-His6) and had a tendency to increase with increasing amount of rHMGB1AB-His6. The cytotoxicity of rHMGB1AB-His6 was compared with poly-l-lysine (PLL, 20Â kDa). As a result, rHMGB1AB-His6 did not show any cytotoxicity to 293 cells, while PLL had a greater cytotoxicity. rHMGB1AB-His6 had the highest transfection efficiency at a 1:40 weight ratio (pDNA/rHMGB1AB-His6). In addition, rHMGB1AB-His6 showed comparable transfection efficiency to PLL. With low cytotoxicity and moderate transfection efficiency, rHMGB1AB-His6 may be useful for delivery of nucleic acids.
Related Topics
Physical Sciences and Engineering
Chemical Engineering
Bioengineering
Authors
Kyunghwa Kim, Jee Seung Han, Jeong Hyun Park, Kyung Soo Ko, Minhyung Lee,