Quantifying structural changes and stoichiometry of protein interactions using size and density profiling |
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Authors: | Marcus Swann Neville Freeman Simon Carrington Gerry Ronan Paul Barrett |
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Institution: | (1) Farfield Sensors Ltd, Unit 51, Salford Business Park, Leslie Hough Way, M6 6AJ Salford, Manchester, UK;(2) ATA Scientific Pty Ltd, Sutherland, PO Box 1005 Sydney, NSW, 1499, Australia |
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Abstract: | Summary The use of Dual Polarisation Interferometry, and emerging analytical biophysical technique, is described for the determination
of the optogeometrical properties (thickness and density) at high resolution of adsorbed protein layers at the solid-liquid
interface. The technique has been used to quantify, in real time and at subatomic resolution, the structural changes occurring
in two well-characterised protein interaction systems, an antibody-antigen interaction and the biotin-streptavidin interaction.
The realtime data obtained on structural changes during the interactions is in excellent agreement with previously reported
X-ray crystallography and neutron reflection data. The precision of the measurements taken was of the order of 0.01 nm with
respect to protein size. The dual-parameter approach also allowed the stoichiometry of both of these interactions to be calculated,
giving values that confirm the current understanding of the interactions. This approach provides detailed insights into the
inherent and subtle link between structural change and function in proteins, to a degree not previously possible through mass
change measurements alone. The technique is expected to find utility in the increasingly important study of protein structure
and function. |
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Keywords: | biophysical techniques conformational change dual polarisation interferometry ligand binding protein function protein interactions protein structure stoichiometry |
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