Identification of lectin counter-receptors on cell membranes by proximity labelling

Gang Wu, Manjula Nagala, Paul R. Crocker (Lead / Corresponding author)

Research output: Contribution to journalArticle

4 Citations (Scopus)
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Abstract

Lectin-glycan interactions play important roles in many biological systems, but the nature of glycoprotein counter-receptors expressed on cell membranes is often poorly understood. To help overcome this problem, we developed a method based on proximity labelling technology. Using a peroxidase-coupled lectin, addition of H 2 O 2 and tyramide-biotin substrates leads to generation of short-range biotin radicals that biotinylate proteins in the immediate vicinity of the bound lectin, which can subsequently be identified. As a proof-of-principle, sialoadhesinhorseradish peroxidase-human IgG1 Fc recombinant protein constructs were precomplexed with anti-Fc antibodies, bound to human erythrocytes and reacted with H 2 O 2 and tyramideSS-biotin. The erythrocyte membrane protein with strongest biotinylation was identified as glycophorin A, in agreement with early studies using lectin overlay and reglycosylation approaches. As a further test of the method, the plant lectin MAL II was conjugated with horseradish peroxidase and used in proximity labelling of human erythrocytes. Glycophorin A was again selectively labelled, which is consistent with previous reports that MAL II has high affinity for glycophorin. This method could be applied to other lectins to identify their membrane counter-receptors.
Original languageEnglish
Pages (from-to)800-805
Number of pages6
JournalGlycobiology
Volume27
Issue number9
Early online date28 Jul 2017
DOIs
Publication statusPublished - 1 Sep 2017

Keywords

  • glycophorin
  • lectin counter-receptor
  • proximity labelling
  • sialoadhesin

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