Blue Devils

Welcome to the Toone group home page. Here you will find descriptions of our various research efforts, references to recent publications, information about the various Departments and Programs at Duke in which students in the Toone lab participate, information about Durham and the Triangle area, and a variety of tools you might find useful in your own research.

Research in the Toone lab surrounds two questions of fundamental importance to the study of biology: what is structural basis of affinity and selectivity in aqueous association events, and what is the structural origin of catalytic efficiency in enzymatic processes, especially mulit-enzyme complexes.  In the course of this work we have developed methodology of universal utility for the evaluation of biological activity at the single molecule level, and facile methods for pattern creation and transfer at sub-100 nm length scales on technologically relevant systems.

Association in aqueous solution is perhaps the most fundamental of biological processes.  Intra- and intermolecular binding controls events as disparate as enzyme-substrate recognition and association, receptor activation and protein folding.  Other association events form membranes and organelles, organize complex multi-enzyme complexes and spatially organize multi-cellular organisms.  Despite the remarkable importance of association to almost every aspect of life, virtually nothing is known regarding the basis of affinity and specificity in aqueous association processes.  The consequences of this gap in our knowledge is profound, leading to an inability to rationally design proteins with pre-determined structures and functions or of small molecules that bind to receptors of known structure with predictable affinities.

The greatest gap in our knowledge base regarding aqueous association surrounds the behavior of water and its interaction with dissolved substrates.  All binding events can be decomposed into two broad processes:  a desolvation event in which the interacting surfaces are removed from water, and an association event, in which the cognate faces of the interacting pair come together and form interactions.  Our understanding of the second component of this parsing is reasonably complete, and the creation of interacting pairs in organic solvent or in the gas phase is by now reasonably straightforward.  On the other hand, the energetic consequences of desolvation are almost completely opaque, and even today our molecular understanding of solvation and desolvation processes is almost totally lacking:  while the hydrophobic effect is well known phenomenonologically its molecular basis is obscure.  We are addressing this vexing fundamental problem through energetic evaluation of the association of synthetic hosts and guests in aqueous and mixed aqueous-organic solvents.  Our experimental tools in this work include theory and isothermal titration microcalorimetry.

Another approach to an understanding of the molecular basis of aqueous association involves a study of additivity in ligand binding.  In this approach, utilized as a fragment-based screening approach in drug discovery, involves the evaluation of the affinity of conceptual ligand fragments and comparing the sum of these affinities to the affinity of the complete ligand.  There is, of course, no reason to assume that simple additivity will hold.  Rather, in the general case any thermodynamic parameter (J) for association of a conceptual ligand AB will be related to the sum of the parameters for the components A and B by the expression:


 where deltaJ is an interaction term that describes the energetic consequences of physical tethering.  An evaluation of interaction enthalpies and entropies will inform the nature of affinity in aqueous solution and better enable both an understanding of the molecular origin of solution thermodynamic properties and the rational design of small molecule ligands for targets of known shape and character.

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In the course of our research we have developed a variety of techniques for both measurement and fabrication.  Recently, we developed an AFM-based approach to the measurement of thermodynamic parameters for ligand binding at the single molecule level.  Briefly, one half of a binding pair is affixed an AFM tip and the other to the corresponding stage.  When the two are brought into physical proximity a complex can form.  Observation of a characteristic force - distance curve during retraction of the stage from the tip reveals the formation of this complex.  Repetition of this approach - retract cycle provides a statistical probability of complex formation between the immobilized pair.  The entire cycle is then repeated, but in the presence of a known quantity of soluble ligand, generating a competitive equilibrium between free and immobilized ligand and resulting in a diminished probability of complex formation.  The difference between the probability of complex formation with and without soluble ligand can be related to the free energy of complex formation for the immobilized ligand - receptor pair.  We are using this methodology to understand the way that various entropic contributors (translational, rotational, configurational) contribute to the thermodynamics of ligand binding.

We have also developed novel approaches to soft lithography and microcontact printing.  Traditional microcontact printing, in which an elastomeric stamp ¡®inked¡¯ with a reactive molecular species, is brought into contact with a surface reactive to the molecular ink.  Pattern is transferred in a diffusive process, as ink diffuses from the stamp to the surface.  The diffusive nature of the process results in pattern ¡®blurring¡¯ near the feature edge, limiting the resolution of traditional microcontact printing to at least hundreds of nanometers.  Our approach to microcontact printing utilizes catalytic stamps that transfer pattern by catalytic modification of an underlying surface.  Our approach provides very high resolution (<100 nm) on a wide range of technologically relevant surfaces, including semiconductor materials and optical materials.

More detailed descriptions of each area can be found in the links in the Project and People page. We urge you to explore our site and to contact us should you have questions regarding any of what you find here.

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