431218 REAL-Time Evaluations of Binding Events Between Streptavidin and Its Ligands

Wednesday, November 11, 2015: 9:50 AM
253A (Salt Palace Convention Center)
Xiao Hu and Cerasela Zoica Dinu, Department of Chemical Engineering, West Virginia University, Morgantown, WV

In the cells, protein-ligand specific recognition involves association and dissociation processes controlled by the affinity of the two partners under chemical harvesting of adenosine triphosphate energy. Fundamental knowledge of selected specific recognition events is currently translated in synthetic environment and widely used in biosensors, immunoassays and diagnosis applications, or in pharmaceutical development. However, in order to advance such fields, one needs to determine the bond lifetime by characterizing the binding efficiency function of the rupture force of the two partners, as well as the energy landscape parameters or the bond and barrier width energy.  

In this study, contact mode analysis of rupture forces between streptavidin and its ligands, namely biotin and anti-streptavidin antibody, were evaluated in real-time using atomic force microscopy (AFM) to provide insights of their relative strength. Our approach used 3-Aminopropyltriethoxysilane (APTS) and glutaraldehyde (GA) to covalently functionalize monolayers of streptavidin with controlled protein packing at glass interfaces, while the AFM tip was used to immobilize biotin/anti-streptavidin antibodies. Analysis revealed that the rupture events between the protein and its individual ligands are characteristics of multi- and single bonds of different strengths and energy landscapes as determined by the structural packing, the symmetry and conformational changes of the individual complexes. Our results also revealed that understanding the importance of the rupture forces between individual protein and its ligand could serve as the first step to protect weak bindings in on-off switches employed in biomedical research applications where specificity and selectivity is foremost sought.

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