Observing and Controlling the Folding Pathway of DNA Origami at the Nanoscale

DNA origami is a powerful method
to fold DNA into rationally designed
nanostructures that holds great promise for bionanotechnology. However,
the folding mechanism has yet to be fully resolved, principally due
to a lack of data with single molecule resolution. To address this
issue, we have investigated in detail, using atomic force microscopy,
the morphological evolution of hundreds of individual rectangular
origamis in solution as a function of temperature. Significant structural
changes were observed between 65 and 55 °C both for folding and
melting, and six structural intermediates were identified. Under standard
conditions, folding was initiated at the edges of the rectangle and
progressed toward the center. Melting occurred through the reverse
pathway until the structures were significantly disrupted but ended
through a different pathway involving out-of-equilibrium chainlike
structures. Increasing the relative concentration of center to edge
staples dramatically modified the folding pathway to a mechanism progressing
from the center toward the edges. These results indicate that the
folding pathway is determined by thermodynamics and suggest a way
of controlling it.