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Genomics Inform > Volume 4(3); 2006 > Article
Unfolded Histidine-Tagged Protein is Immobilized to Nitrilotriacetic Acid-Nickel Beads, But Not the Nickel-Coated Glass Slide.
Minho Cho, Sunyoung Ahn, Heonyong Park
Department of Molecular Biology & Institute of Nanosensor and Biotechnology, BK21 Graduate Program for RNA biology, Dankook University, Seoul, Korea. heonyong@dankook.ac.kr
Abstract
The adsorption of proteins on the surface of glass slides is essential for construction of protein chips. Previously, we prepared a nickel-coated plate by the spin-coating method for immobilization of His-tagged proteins. In order to know whether the structural factor is responsible for the immobilization of His-tagged proteins to the nickel-coated glass slide, we executed a series of experiments. First we purified a His-tagged protein after expressing the vector in E. coli BL21 (DE3). Then we obtained the unfolding curve for the His-tagged protein by using guanidine hydrochloride. Fractions unfolded were monitored by internal fluorescence spectroscopy. The delta G(H20) for unfolding was 2.27 kcalmol +/- 0.52. Then we tested if unfolded His-tagged proteins can be adsorbed to the nickel-coated plate, comparing with Ni2+ -NTA (nitrilotriacetic acid) beads. Whereas unfolded His-tagged proteins were adsorbed to Ni2+ -NTA beads, they did not bind to the nickel-coated plate. In conclusion, a structural factor is likely to be an important factor for constructing the protein chips, when His-tagged proteins will immobilize to the nickel-coated slides.
Keywords: his-tagged protein; nickel-coated glass slides; protein chip; unfolding
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