Forming, Counting, and Breaking Individual Biological Bonds: Applications for Drug Delivery and Sing

Todd Sulchek Staff Scientist
Biosecurity and Nanosciences Laboratory
Lawrence Livermore National Laboratory Livermore, CA Protein molecules commonly operate in complexes to perform their function. For example, cell surface receptors often cluster at the site of complementary ligands so as to efficiently transduce binding. A special case of improved functionality through complexed protein binding is demonstrated in a new class of therapeutics in which monovalent antibody binding elements are combined to form multivalent complexes that dramatically increase drug specificity and residency time. However, traditional methods of analysis cannot directly measure the bond lifetime of drug molecules that can bind with a distribution of valencies. Therefore, a single molecule binding assay is illuminating.
We have developed a method using single molecule dynamic force spectroscopy to determine the binding strength of antibody-protein complexes as a function of binding valency in a direct and simple measurement. We used the atomic force microscope (AFM) to measure the force required to rupture a single complex formed by the MUC1 protein, a cancer indicator, and therapeutic antibodies that target MUC1.
We show for the first time that the valency of stochastic, multivalent bond formation can be distinguished with a "molecular counter" in the form of a soft polymer linker. As a result, we independently measure both the valency and the composite bond strength for the interaction. The effective bond lifetime rises dramatically with the number of molecular bonds, from several minutes for a single antibody-antigen bond to many days for three antibody-antigen bonds. Moreover, our results support the theoretical prediction for unbinding dynamics of multiple parallel bonds. We furthermore describe current experiments in which we study cell signal transduction using controlled delivery of protein stimuli.

Location: Stauffer Science Lecture Hall (SLH) Rm 100

Audiences: Everyone Is Invited

Contact: April Mundy