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Orientation of transferrin molecules on mica and amino - derivatized mica studied with SPM using sugar specific lectins

Magnus Bergkvist*, a, Jan Carlssonb,c, and Sven Oscarsson a

aDept. Chemical Engineering, Mälardalen University, Box 325, S-631 05, Eskilstuna, Sweden
bPharmacia & Upjohn Diagnostics AB, S-75182 Uppsala, Sweden
cSurface Biotechnology Center, Uppsala University, Box577, S-75123, Uppsala

This is an abstract for a presentation given at the
Sixth Foresight Conference on Molecular Nanotechnology.
There will be a link from here to the full article when it is available on the web.

 

Knowledge about the adsorption and orientation of biomolecules to surfaces is of importance for creation of complex molecular structures on surfaces. With Atomic Force Microscopy it is possible to increase the resolution and study individual molecules and their orientation to surfaces. In this study we compare the orientation of Human-Serum (hS - ) Transferrin upon adsorption to mica and Aminopropylmethyldietoxysilane (APDES)-mica surfaces with Atomic Force Microscopy using sugar-specific elderberry bark lectin as a molecular probe. Since hS-Transferrin has two branched sugar chains relatively close to each other in its C-terminal domain, it is a perfect model protein for demonstrating how a lectin molecule can be used as a probe of protein orientation and adsorption properties. Adsorbed molecules were analyzed regarding height and area, which were obtained through the nanoscope software bearing-analysis feature. The volume of the molecules was calculated by approximating the shape of a rectangle and multiplying area times height, which results in a distribution of volumes. Transferrin adsorbed to both mica and APDES-mica surfaces yields a volume distribution peak within 225-675 nm3 (Fig1, Fig2) and one population when plotting area vs. height. When adding lectin to the tranferrin, the transferrin population decreases and the binding of lectin generates a larger volume distribution and also a new population in the area vs. height diagram. In oreder to show that this is a specific interaction, galactose, which inhibits the lectin at higher concentrations, was added. In doing so the larger distribution peak disappeared and just the original transferrin peak at 225-675 nm3 was present. When comparing the different results for mica and APDES mica it is clear that there is a different orientation/adsorption to the two surfaces. On mica lectin binds to all transferrin (Fig1), while on APDES-mica only 40 % is capable of binding (Fig2). This indicates that the C-terminal domain with the sugar chains is oriented in such a way that lectin binding is reduced.


*Corresponding Address:
Magnus Bergkvist
Dept. of  Biology and Chemical Engineering, Gammla tullgatan2, S-63125, Eskilstuna, Sweden
Phone:+46-16-153461, Fax:+46-16-153710
Email: magnus.bergkvist@mdh.se, Web: http://www.mdh.se/organisation/ibk/forskning/index.htm



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