Entirely enzymatic nanofabrication of DNA-protein conjugates

Nucleic Acids Res. 2017 Oct 13;45(18):e160. doi: 10.1093/nar/gkx707.

Abstract

While proteins are highly biochemically competent, DNA offers the ability to program, both reactions and the assembly of nanostructures, with a control that is unprecedented by any other molecule. Their joining: DNA-protein conjugates - offer the ability to combine the programmability of DNA with the competence of proteins to form novel tools enabling exquisite molecular control and the highest biological activity in one structure. However, in order for tools like these to become viable for biological applications, their production must be scalable, and an entirely enzymatic process is one way to achieve this. Here, we present a step in this direction: enzymatic production of DNA-protein conjugates using a new self-labeling tag derived from a truncated VirD2 protein of Agrobacterium tumefaciens. Using our previously reported MOSIC method for enzymatic ssDNA oligo production, we outline a pipeline for protein-DNA conjugates without the need for any synthetic chemistry in a one-pot reaction. Further, we validate HER2 staining using a completely enzymatically produced probe, enable the decoration of cell membranes and control of genetic expression. Establishing a method where protein-DNA conjugates can be made entirely using biological or enzymatic processing, opens a path to harvest these structures directly from bacteria and ultimately in-vivo assembly.

MeSH terms

  • Agrobacterium tumefaciens / enzymology*
  • Agrobacterium tumefaciens / genetics
  • Agrobacterium tumefaciens / metabolism
  • Bacterial Proteins / metabolism*
  • Cells, Cultured
  • DNA / metabolism*
  • HEK293 Cells
  • Humans
  • Metabolic Engineering / methods
  • Nanostructures / chemistry*
  • Nanotechnology / methods*
  • Protein Binding
  • Synthetic Biology / methods

Substances

  • Bacterial Proteins
  • DNA