Covalent protein-oligonucleotide conjugates by copper-free click reaction.

Published

Journal Article

Covalent protein-oligodeoxynucleotide (protein-ODN) conjugates are useful in a number of biological applications, but synthesizing discrete conjugates-where the connection between the two components is at a defined location in both the protein and the ODN-under mild conditions with significant yield can be a challenge. In this article, we demonstrate a strategy for synthesizing discrete protein-ODN conjugates using strain-promoted azide-alkyne [3+2] cycloaddition (SPAAC, a copper-free 'click' reaction). Azide-functionalized proteins, prepared by enzymatic prenylation of C-terminal CVIA tags with synthetic azidoprenyl diphosphates, were 'clicked' to ODNs that had been modified with a strained dibenzocyclooctyne (DIBO-ODN). The resulting protein-ODN conjugates were purified and characterized by size-exclusion chromatography and gel electrophoresis. We find that the yields and reaction times of the SPAAC bioconjugation reactions are comparable to those previously reported for copper-catalyzed azide-alkyne [3+2] cycloaddition (CuAAC) bioconjugation, but require no catalyst. The same SPAAC chemistry was used to immobilize azide-modified proteins onto surfaces, using surface-bound DIBO-ODN as a heterobifunctional linker. Cu-free click bioconjugation of proteins to ODNs is a simple and versatile alternative to Cu-catalyzed click methods.

Full Text

Duke Authors

Cited Authors

  • Khatwani, SL; Kang, JS; Mullen, DG; Hast, MA; Beese, LS; Distefano, MD; Taton, TA

Published Date

  • July 15, 2012

Published In

Volume / Issue

  • 20 / 14

Start / End Page

  • 4532 - 4539

PubMed ID

  • 22682299

Pubmed Central ID

  • 22682299

Electronic International Standard Serial Number (EISSN)

  • 1464-3391

Digital Object Identifier (DOI)

  • 10.1016/j.bmc.2012.05.017

Language

  • eng

Conference Location

  • England