De novo resonance assignment of the transmembrane domain of LR11/SorLA in E. coli membranes

Xiaoyan Ding, Riqiang Fu, Fang Tian

Research output: Contribution to journalArticle

Abstract

Membrane proteins perform many important cellular functions. Historically, structural studies of these proteins have been conducted in detergent preparations and synthetic lipid bilayers. More recently, magic-angle-spinning (MAS) solid-state NMR has been employed to analyze membrane proteins in native membrane environments, but resonance assignments with this technique remain challenging due to limited spectral resolution and high resonance degeneracy. To tackle this issue, we combine reverse labeling of amino acids, frequency-selective dipolar dephasing, and NMR difference spectroscopy. These methods have resulted in nearly complete resonance assignments of the transmembrane domain of human LR11 (SorLA) protein in E. coli membranes. To reduce background signals from E. coli lipids and proteins and improve spectral sensitivity, we effectively utilize amylose affinity chromatography to prepare membrane vesicles when MBP is included as a fusion partner in the expression construct.

Original languageEnglish (US)
Article number106639
JournalJournal of Magnetic Resonance
Volume310
DOIs
StatePublished - Jan 2020

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Escherichia coli
membranes
proteins
Membranes
Membrane Proteins
Nuclear magnetic resonance
Affinity chromatography
Amylose
Magic angle spinning
Lipid bilayers
Escherichia coli Proteins
Spectral resolution
Lipid Bilayers
lipids
Affinity Chromatography
Detergents
Labeling
Proteins
Magnetic Resonance Spectroscopy
Fusion reactions

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Nuclear and High Energy Physics
  • Condensed Matter Physics

Cite this

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title = "De novo resonance assignment of the transmembrane domain of LR11/SorLA in E. coli membranes",
abstract = "Membrane proteins perform many important cellular functions. Historically, structural studies of these proteins have been conducted in detergent preparations and synthetic lipid bilayers. More recently, magic-angle-spinning (MAS) solid-state NMR has been employed to analyze membrane proteins in native membrane environments, but resonance assignments with this technique remain challenging due to limited spectral resolution and high resonance degeneracy. To tackle this issue, we combine reverse labeling of amino acids, frequency-selective dipolar dephasing, and NMR difference spectroscopy. These methods have resulted in nearly complete resonance assignments of the transmembrane domain of human LR11 (SorLA) protein in E. coli membranes. To reduce background signals from E. coli lipids and proteins and improve spectral sensitivity, we effectively utilize amylose affinity chromatography to prepare membrane vesicles when MBP is included as a fusion partner in the expression construct.",
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De novo resonance assignment of the transmembrane domain of LR11/SorLA in E. coli membranes. / Ding, Xiaoyan; Fu, Riqiang; Tian, Fang.

In: Journal of Magnetic Resonance, Vol. 310, 106639, 01.2020.

Research output: Contribution to journalArticle

TY - JOUR

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