The His-tag as a decoy modulating preferred orientation in cryoEM

Raquel Bromberg, Kai Cai, Yirui Guo, Daniel Plymire, Tabitha Emde, Maciej Puzio, Dominika Borek, Zbyszek Otwinowski

Research output: Contribution to journalArticlepeer-review

1 Scopus citations


The His-tag is a widely used affinity tag that facilitates purification by means of affinity chromatography of recombinant proteins for functional and structural studies. We show here that His-tag presence affects how coproheme decarboxylase interacts with the air-water interface during grid preparation for cryoEM. Depending on His-tag presence or absence, we observe significant changes in patterns of preferred orientation. Our analysis of particle orientations suggests that His-tag presence can mask the hydrophobic and hydrophilic patches on a protein’s surface that mediate the interactions with the air-water interface, while the hydrophobic linker between a His-tag and the coding sequence of the protein may enhance other interactions with the air-water interface. Our observations suggest that tagging, including rational design of the linkers between an affinity tag and a protein of interest, offer a promising approach to modulating interactions with the air-water interface.

Original languageEnglish (US)
Article number912072
JournalFrontiers in Molecular Biosciences
StatePublished - Oct 17 2022


  • air-water interface
  • AWI
  • cryo-electron microscopy single particle reconstruction
  • cryoEM SPR
  • His-tag
  • preferred orientation
  • visualization

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology (miscellaneous)


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