Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags

D. Wasserberg, Jordi Cabanas-Danés, Jord Prangsma, Shane O'Mahony, Pierre Andre Cazade, E. Tromp, Christian Blum, Damien Thompson, Jurriaan Huskens, Vinod Subramaniam, Pascal Jonkheijm

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Abstract

We report oriented immobilization of proteins using the standard hexahistidine (His6)-Ni2+:NTA (nitrilotriacetic acid) methodology, which we systematically tuned to give control of surface coverage. Fluorescence microscopy and surface plasmon resonance measurements of self-assembled monolayers (SAMs) of red fluorescent proteins (TagRFP) showed that binding strength increased by 1 order of magnitude for each additional His6-tag on the TagRFP proteins. All TagRFP variants with His6-tags located on only one side of the barrel-shaped protein yielded a 1.5 times higher surface coverage compared to variants with His6-tags on opposite sides of the so-called β-barrel. Time-resolved fluorescence anisotropy measurements supported by polarized infrared spectroscopy verified that the orientation (and thus coverage and functionality) of proteins on surfaces can be controlled by strategic placement of a His6-tag on the protein. Molecular dynamics simulations show how the differently tagged proteins reside at the surface in "end-on" and "side-on" orientations with each His6-tag contributing to binding. Also, not every dihistidine subunit in a given His6-tag forms a full coordination bond with the Ni2+:NTA SAMs, which varied with the position of the His6-tag on the protein. At equal valency but different tag positions on the protein, differences in binding were caused by probing for Ni2+:NTA moieties and by additional electrostatic interactions between different fractions of the β-barrel structure and charged NTA moieties. Potential of mean force calculations indicate there is no specific single-protein interaction mode that provides a clear preferential surface orientation, suggesting that the experimentally measured preference for the end-on orientation is a supra-protein, not a single-protein, effect.

Original languageEnglish
Pages (from-to)9068-9083
Number of pages16
JournalACS nano
Volume11
Issue number9
DOIs
Publication statusPublished - 26 Sep 2017

Fingerprint

His-His-His-His-His-His
histidine
Histidine
Membrane Proteins
proteins
Proteins
Self assembled monolayers
Nitrilotriacetic Acid
fluorescence
Fluorescence microscopy
Surface plasmon resonance
Coulomb interactions

Keywords

  • molecular dynamics simulations
  • monolayers
  • multivalency
  • protein immobilization
  • self-assembly

Cite this

Wasserberg, D., Cabanas-Danés, J., Prangsma, J., O'Mahony, S., Cazade, P. A., Tromp, E., ... Jonkheijm, P. (2017). Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags. ACS nano, 11(9), 9068-9083. https://doi.org/10.1021/acsnano.7b03717
Wasserberg, D. ; Cabanas-Danés, Jordi ; Prangsma, Jord ; O'Mahony, Shane ; Cazade, Pierre Andre ; Tromp, E. ; Blum, Christian ; Thompson, Damien ; Huskens, Jurriaan ; Subramaniam, Vinod ; Jonkheijm, Pascal. / Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags. In: ACS nano. 2017 ; Vol. 11, No. 9. pp. 9068-9083.
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Wasserberg, D, Cabanas-Danés, J, Prangsma, J, O'Mahony, S, Cazade, PA, Tromp, E, Blum, C, Thompson, D, Huskens, J, Subramaniam, V & Jonkheijm, P 2017, 'Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags' ACS nano, vol. 11, no. 9, pp. 9068-9083. https://doi.org/10.1021/acsnano.7b03717

Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags. / Wasserberg, D.; Cabanas-Danés, Jordi; Prangsma, Jord; O'Mahony, Shane; Cazade, Pierre Andre; Tromp, E.; Blum, Christian; Thompson, Damien; Huskens, Jurriaan; Subramaniam, Vinod; Jonkheijm, Pascal.

In: ACS nano, Vol. 11, No. 9, 26.09.2017, p. 9068-9083.

Research output: Contribution to journalArticleAcademicpeer-review

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AU - Wasserberg, D.

AU - Cabanas-Danés, Jordi

AU - Prangsma, Jord

AU - O'Mahony, Shane

AU - Cazade, Pierre Andre

AU - Tromp, E.

AU - Blum, Christian

AU - Thompson, Damien

AU - Huskens, Jurriaan

AU - Subramaniam, Vinod

AU - Jonkheijm, Pascal

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Wasserberg D, Cabanas-Danés J, Prangsma J, O'Mahony S, Cazade PA, Tromp E et al. Controlling Protein Surface Orientation by Strategic Placement of Oligo-Histidine Tags. ACS nano. 2017 Sep 26;11(9):9068-9083. https://doi.org/10.1021/acsnano.7b03717