Project: Research project

Project Details


Protein-mediated molecular adhesion: AFM studies.

Numerous cellular processes including cell adhesion and cell migration are
mediated by the transient interaction of adhesion molecules. Often, as in
the case cell migration, there is a delicate balance between cell
attachment and cell de-attachment. The strength of cell attachment is
determined by the number of intermolecular bonds formed between membrane-
bound receptors and their ligands on the opposing surface and the de-
attachment force of the individual bonds. Whereas the number of bonds
formed at the interface is closely related to the association constant of
the protein-ligand interaction, the rupture force of these bonds is poorly
understood and is the main focus of this proposal.

The long-term objective of this proposed research is to achieve a
fundamental understanding of the mechanisms involved in molecular adhesion.
The current proposal examines the intermolecular forces of protein-ligand
interaction in two model systems, the streptavidin-biotin pair and an
antibody-antigen pair. In general, protein-ligand bonds are noncovalent
and will spontaneously break by thermal agitation given sufficient time.
The dissociation lifetime of the protein-ligand bond is accelerated and can
be studied by applying an external force across the bond with an Atomic
Force Microscope (AFM). Comprehensive measurements of force-life time
relationships derived from AFM-induced separation of the streptavidin-
biotin bond will be used to reveal the dissociation pathway and possible
intermediate states of the complex. AFM measurements of mutagenized
streptavidin will be used to identify key amino acid determinants
responsible for adhesion to biotin. In contrast to the resilience of the
streptavidin-biotin interaction, the binding of fluorescein to 4-4-20, an
anti-fluorescyl antibody, can be thermodynamically manipulated by small
changes in temperature, pH, and solvent system. The effects of these
perturbations on the dissociation constant and binding enthalpy will be
studied and correlated to force measurements to determine the reaction
pathway of 4-4-20:fluorescein dissociation beyond the point of initial bond
rupture. Together, these experiments will contribute to establishing a
conceptual framework for understanding protein-mediated molecular adhesion.
Effective start/end date5/1/974/30/03


  • National Institute of General Medical Sciences: $10,688.00
  • National Institute of General Medical Sciences
  • National Institute of General Medical Sciences: $150,182.00
  • National Institute of General Medical Sciences
  • National Institute of General Medical Sciences
  • National Institute of General Medical Sciences: $154,449.00


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