Unfolded Histidine-Tagged Protein is Immobilized to Nitrilotriacetic Acid-Nickel Beads, But Not the Nickel-Coated Glass Slide.
- Author:
Minho CHO
1
;
Sunyoung AHN
;
Heonyong PARK
Author Information
1. Department of Molecular Biology & Institute of Nanosensor and Biotechnology, BK21 Graduate Program for RNA biology, Dankook University, Seoul, Korea. heonyong@dankook.ac.kr
- Publication Type:Original Article
- Keywords:
his-tagged protein;
nickel-coated glass slides;
protein chip;
unfolding
- MeSH:
Adsorption;
Fibrinogen;
Glass*;
Guanidine;
Immobilization;
Protein Array Analysis;
Spectrometry, Fluorescence
- From:Genomics & Informatics
2006;4(3):133-136
- CountryRepublic of Korea
- Language:English
-
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.
