Journal of Biomolecular NMR

, 51:173 | Cite as

Topology and immersion depth of an integral membrane protein by paramagnetic rates from dissolved oxygen

  • M. Sameer Al-Abdul-Wahid
  • Raffaello Verardi
  • Gianluigi Veglia
  • R. Scott Prosser
Article
First Online:
Received:
Accepted:

Abstract

In studies of membrane proteins, knowledge of protein topology can provide useful insight into both structure and function. In this work, we present a solution NMR method for the measurement the tilt angle and average immersion depth of alpha helices in membrane proteins, from analysis of the paramagnetic relaxation rate enhancements arising from dissolved oxygen. No modification to the micelle or protein is necessary, and the topology of both transmembrane and amphipathic helices are readily determined. We apply this method to the measure the topology of a monomeric mutant of phospholamban (AFA-PLN), a 52-residue membrane protein containing both an amphipathic and a transmembrane alpha helix. In dodecylphosphocholine micelles, the amphipathic helix of AFA-PLN was found to have a tilt angle of 87° ± 1° and an average immersion depth of 13.2 Å. The transmembrane helix was found to have an average immersion depth of 5.4 Å, indicating residues 41 and 42 are closest to the micelle centre. The resolution of paramagnetic relaxation rate enhancements from dissolved oxygen compares favourably to those from Ni (II), a hydrophilic paramagnetic species.

Keywords

Membrane protein immersion depth Membrane protein topology Paramagnetic relaxation rate enhancements Solution state NMR of membrane proteins 

Abbreviations

AFA-PLN

Phospholamban mutant C36A/C41F/C46A

DOPC

1,2-dioleoyl-glycero-3-sn-phosphocholine

DOPE

Dioleylphosphoethanolamine

EPR

Electron paramagnetic resonance

HSQC

Heteronuclear single quantum correlation

NOE

Nuclear overhauser effect

PISEMA

Polarization inversion spin exchange at magic angle

PLN, wt-PLN

Phospholamban

PRE

Paramagnetic relaxation enhancement

RDC

Residual dipolar coupling

SDS

Sodium dodecyl sulfate

SERCA

Sarco(endo)plasmic reticulum calcium ATPase

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Notes

Acknowledgments

RSP acknowledges NSERC, and the Ontario government for financial support through the NSERC discovery and Provincial Research Excellence Award (PREA) programs. This work was in part supported by the National Institute of Health (GM64742) to GV.

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • M. Sameer Al-Abdul-Wahid
    • 1
  • Raffaello Verardi
    • 2
    • 3
  • Gianluigi Veglia
    • 2
    • 3
  • R. Scott Prosser
    • 1
    • 4
  1. 1.Department of ChemistryUniversity of Toronto MississaugaMississaugaCanada
  2. 2.Department of ChemistryUniversity of MinnesotaMinneapolisUSA
  3. 3.Department of Biochemistry, Molecular Biology, and BiophysicsUniversity of MinnesotaMinneapolisUSA
  4. 4.Department of BiochemistryUniversity of TorontoTorontoCanada

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