Organizing Enzymes on Self-Assembled Protein Cages for Cascade Reactions.

Cells use self-assembled biomaterials such as lipid membranes or proteinaceous shells to coordinate thousands of reactions that simultaneously take place within crowded spaces. However, mimicking such spatial organization for synthetic applications in engineered systems remains a challenge, resulting in inferior catalytic efficiency. In this work, we show that protein cages as an ideal scaffold to organize enzymes to enhance cascade reactions both in vitro and in living cells. We demonstrate that not only enzyme-enzyme distance but also the improved Km value contribute to the enhanced reaction rate of cascade reactions. Three sequential enzymes for lycopene biosynthesis have been co-localized on the exterior of the engineered protein cages in Escherichia coli, leading to an 8.5-fold increase of lycopene production by streamlining metabolic flux towards its biosynthesis. This versatile system offers a powerful tool to achieve enzyme spatial organization for broad applications in biocatalysis.

Contact-coupled impact of slender rods: analysis and experimental validation.

To validate models of contact mechanics in low speed structural impact, slender rods were impacted in a drop tower, and measurements of the contact and vibration were compared to analytical and finite element (FE) models. The contact area was recorded using a novel thin-film transfer technique, and the contact duration was measured using electrical continuity. Strain gages recorded the vibratory strain in one rod, and a laser Doppler vibrometer measured speed. The experiment was modeled analytically on a one-dimensional spatial domain using a quasi-static Hertzian contact law and a system of delay differential equations. The three-dimensional FE model used hexahedral elements, a penalty contact algorithm, and explicit time integration. A small submodel taken from the initial global FE model economically refined the analysis in the small contact region. Measured contact areas were within 6% of both models' predictions, peak speeds within 2%, cyclic strains within 12 µÎµ (RMS value), and contact durations within 2 µs. The global FE model and the measurements revealed small disturbances, not predicted by the analytical model, believed to be caused by interactions of the non-planar stress wavefront with the rod's ends. The accuracy of the predictions for this simple test, as well as the versatility of the diagnostic tools, validates the theoretical and computational models, corroborates instrument calibration, and establishes confidence that the same methods may be used in experimental and computational study of contact mechanics during impact of more complicated structures. Recommendations are made for applying the methods to a particular biomechanical problem: the edge-loading of a loose prosthetic hip joint which can lead to premature wear and prosthesis failure.