| AVS 65th International Symposium & Exhibition | |
| Biomaterial Interfaces Division | Tuesday Sessions |
| Session BI+AS+IPF+NS-TuA |
| Session: | IoT Session: Biofabrication, Bioanalytics, Biosensors and Diagnostics and Flash Networking Session |
| Presenter: | Nourin Alsharif, Boston University |
| Authors: | N. Alsharif, Boston University T. Lawton, Natick Soldier Research, Development and Engineering Center J. Uzarski, Natick Soldier Research, Development and Engineering Center K.A. Brown, Boston University |
| Correspondent: | Click to Email |
Many of the exceptional properties of natural materials (e.g. fracture toughness of bones, strength to weight ratio of bamboo) can be attributed to their structural hierarchy, which originates, in part, from the nanoscale organization of the enzymes that synthesize these materials. In order to best utilize such enzymes ex vivo to grow engineered biomaterials, the role of this multiscale organization must be understood. Here, we report a novel strategy for studying the activity of arrangements of enzymes within a multifunctional material in a high throughput manner. In particular, we use top-down patterning techniques in conjunction with small molecule self-assembly to designate enzyme-binding regions amidst a non-binding, hydrophobic background. Key to this experimental scheme is the parallel nature of both the fabrication and the characterization processes that enable the efficient study of many geometric parameters of the enzyme-binding features. These parameters include, (1) feature size, (2) density of enzyme within each feature, and (3) distance between features. This level of control can in principle allow us to separate effects of reaction kinetics and substrate diffusion. Two strategies have been explored for the immobilization of enzymes including click chemistry to non-natural amino acids and binding to poly-histidine affinity tags. Top-down lithography and enzyme assembly were verified using a variety of surface characterization techniques including atomic force microscopy, X-ray photoelectron spectroscopy, infrared spectroscopy, spectroscopic ellipsometry, and contact angle goniometry. Initially, this high throughput paradigm is used to develop a fluorimetric assay to quantify the activity of surface-bound enzymes as a function of their spatial organization. Together with the widespread utilization of high throughput techniques in synthetic biology, the ability to study spatial organization in a rapid fashion is expected to dramatically improve ex vivo applications of enzymes.