Abstract
We have devised a supramolecular edifice involving His-tagged protein A and antibodies to yield surface immobilized, uniformly oriented, IgG-type, antibody layers with Fab fragments exposed off an electrode surface. We demonstrate here that we can affect the conformation of IgGs, likely pushing/pulling electrostatically Fab fragments towards/from the electrode surface. A potential difference between electrode and solution acts on IgGs’ charged aminoacids modulating the accessibility of the specific recognition regions of Fab fragments by antigens in solution. Consequently, antibody-antigen affinity is affected by the sign of the applied potential: a positive potential enables an effective capture of antigens; a negative one pulls the fragments towards the electrode, where steric hindrance caused by neighboring molecules largely hampers the capture of antigens. Different experimental techniques (electrochemical quartz crystal microbalance, electrochemical impedance spectroscopy, fluorescence confocal microscopy and electrochemical atomic force spectroscopy) were used to evaluate binding kinetics, surface coverage, effect of the applied electric field on IgGs, and role of charged residues on the phenomenon described. These findings expand the concept of electrical control of biological reactions and can be used to gate electrically specific recognition reactions with impact in biosensors, bioactuators, smart biodevices, nanomedicine, and fundamental studies related to chemical reaction kinetics.
Highlights
The possibility of acting on biomolecules using an applied electric field is at the basis of many methods and approaches adopted in different contexts such as bioanalysis, diagnosis and therapy, nanobiotechnology, and molecular electronics[1,2,3]
IgGs’ positive charges from the surface, pushing Fab fragments towards the solution bulk; antigen binding from solution is favored. (b) A negative substrate potential attracts IgGs’ positively charged residues, pulling Fab fragments towards the electrode surface; in this case, antigen binding is made less probable due to a decreased accessibility of specific recognition sites by antigens in solution
The first step was that of developing a surface immobilization chemistry for antibodies on a gold electrode surface that enabled their preferential orientation, that of their Fab fragments, towards the solution covering the electrode
Summary
The possibility of acting on biomolecules using an applied electric field is at the basis of many methods and approaches adopted in different contexts such as bioanalysis, diagnosis and therapy, nanobiotechnology, and molecular electronics[1,2,3]. This possibility stems from the fact that, in physiologic conditions, biomolecules possess net electric charges and generally have quite complex charge distributions[4], which make them sensitive to the presence of external electric fields.
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