3D Structure of DNA-Protein 2D Lattice by Cryo-Electron Tomography

Programmable and self-assembled two-dimensional (2D) protein lattices hold tremendous potential in the realms of biological crystalline devices and nanoscale catalysis, signifying a pivotal advancement from the fundamental building blocks towards intricate three-dimensional (3D) crystal structures. However, the creation of high-order lattices remains challenging, partly due to the lack of a tool to precisely analyze the 3D structure of each building block within short-range-ordered crystals without averaging. In this study, we leveraged advances in cryogenic electron tomography (cryo-ET) and the individual-particle electron tomography (IPET) technique to determine the 3D structure of each individual building block within short-range-ordered 2D protein-DNA origami lattices. The building block features a DNA origami octahedral cage, composed of 12 DNA helical bundles, and encapsulates ferritin molecules. Our quantitative analysis of the variations of conformations and orientational of these distinctive and asymmetric building blocks showed that ferritin loading exerted a negligible influence on the lattice unit-cell parameters. However, the flexibility of the blocks themselves and their connectors significantly affected lattice fluctuations. These findings were further validated by molecular dynamics (MD) simulations and juxtaposed with small-angle scattering (SAXS) data. This research offers an innovative approach to troubleshooting high-order crystallization by evaluating the crystal structure through the lens of the 3D structure of each unit cell.