Collaborative Research: A Stacked Plasmonic Nanopore for Tether-Free Stretching and Label-Free Sensing of hSTf Dynamics and Complex Formation at Ultra-Low Concentrations

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Fundamental knowledge of protein structures and their dynamic responses to stimuli or other molecules is important for many applications, including medical diagnosis and therapy. This research aims to develop a highly sensitive approach for studying the human serum transferrin protein (hSTf), which is a vital iron carrier in blood and of clinical importance. The sensing technique would allow differentiation of the free hSTf protein from the iron-bound protein and evaluation of iron deficiency or iron overload from very small blood samples. Successful development of this sensor would also enable profiling of a wide range of other proteins and biological molecules, e.g., DNA. This project offers excellent opportunities for interdisciplinary research training as it combines biochemistry, nanoengineering, photonics, and electrical engineering. The outreach efforts to K-12 schools through various programs at the Southern Methodist University and the University of Texas at Arlington help to inspire more students to pursue science, technology, engineering and mathematics (STEM) degrees.The stacked plasmonic nanosensor is based on the self-induced back-action (SIBA) actuated nanopore electrophoresis (SANE) sensing concept. The stacked nanopores are uniquely designed to enable 1) controlled trapping, releasing, and recapturing of proteins or the substrate-bound protein complexes, 2) transient deformation of the biological molecules, which can be induced by thermal effect or a combination of optical and electrical techniques, and 3) study of their deformation dynamics. The SANE concept implemented in the stacked nanopore sensor allows investigation of protein interactions at concentrations 1000-fold below the equilibrium dissociation constant in bulk solution, making this technique ultra-sensitive. An important aim of this research is the study of the properties of free-hSTf protein and the iron-bound protein complex using the SANE sensor. Optical signature profiles are established for each of the species to enable selective admission of bound complexes over unbound proteins in a mixed solution to the underlying pore. It uses symmetric (VCapture = VRecapture), followed by asymmetric (VCapture ≠ VRecapture) voltage conditions to facilitate the investigation of the strength and kinetic parameters associated with protein-substrate binding, protein relaxation times, and whether voltage-induced protein unfolding is reversible or not.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.