Sequence-Directed Covalent Protein-DNA Linkages in a Single Step Using HUH-Tags

Klaus N. Lovendahl, Amanda N. Hayward, Wendy R Gordon

Research output: Contribution to journalArticlepeer-review

22 Scopus citations

Abstract

We present a robust strategy to covalently link proteins and DNA using HUH-endonuclease domains as fusion partners (HUH-tags). We show that HUH-tags react robustly with specific sequences of unmodified single-stranded DNA, and we have identified five tags that react orthogonally with distinct DNA sequences. We demonstrate the versatility of HUH-tags as fusion partners in Cas9-mediated gene editing and the construction of doubly DNA-tethered proteins for single-molecule studies. Finally we demonstrate application to cellular imaging in live and fixed cells.

Original languageEnglish (US)
Pages (from-to)7030-7035
Number of pages6
JournalJournal of the American Chemical Society
Volume139
Issue number20
DOIs
StatePublished - May 24 2017

Bibliographical note

Funding Information:
This study was supported by an NIH NIGMS R35 grant to W.R.G. (1R35GM119483), funds to W.R.G. from the Pew Biomedical Scholar Program (29634), and UMN startup funds. We would like to thank Peter Gordon for the kind gift of inducible GFP cell lines. We also would like to thank Eric Hendrickson and Brian Ruis for helpful discussions about the Cas9 experiments. Typhoon FLA9500 images were collected at the University of Minnesota Imaging Center.

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