A new fungal lectin recognizing α(1-6)-linked fucose in the N-glycan

Yasuo Oda, Tsutomu Senaha, Yuuki Matsuno, Kazuki Nakajima, Ryousuke Naka, Mitsuhiro Kinoshita, Eiko Honda, Itaru Furuta, Kazuaki Kakehi

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Abstract

In this report, we describe a new lectin from the fungus Rhizopus stolonifer that agglutinates rabbit red blood cells. Agglutinating activity was detected in the extract of mycelium-forming spores cultured on agar plates but not in the mycelium-forming no spores from liquid medium. This lectin, which we designated R. stolonifer lectin (RSL), was isolated by affinity chromatography with porcine stomach mucin-Sepharose. SDS-polyacrylamide gel electrophoresis and mass spectral analysis showed that RSL is ∼4.5 kDa, whereas gel filtration indicated a mass of 28 kDa. This indicates that the lectin is a hexamer of noncovalently associated RSL monomers. RSL activity was very stable, since it was insensitive to heat treatment at 70 °C for 10 min. Analysis of RSL binding specificity by both microtiter plate and precipitation assays showed that N-glycans with L-fucose linked to the reducing terminal GlcNAc residues are the most potent inhibitors of RSL binding, whereas N-glycans without α(1-6)-linked fucose residues are ∼100-fold weaker inhibitors of binding. Oligosaccharides with α(1-2, -3, and -4) linkages showed no inhibition of binding in these assays. In a mirror resonance biosensor assay, high affinity binding was observed between RSL and the glycopeptide of bovine γ-globulin, which has N-glycans with α(1-6)-linked fucose residues. However, RSL showed only a weak interaction with the glycopeptide of quail ovomucoid, which lacks fucose residues. Finally, capillary affinity electrophoresis studies indicated that RSL binds strongly to N-glycans with α(1-6)-linked fucose residues. Together, these results show that RSL recognizes the core structure of N-glycans with α(1-6)-linked L-fucose residues. This specificity could make RSL a valuable tool for glycobiological studies.

Original languageEnglish
Pages (from-to)32439-32447
Number of pages9
JournalJournal of Biological Chemistry
Volume278
Issue number34
DOIs
Publication statusPublished - 22-08-2003

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All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Oda, Y., Senaha, T., Matsuno, Y., Nakajima, K., Naka, R., Kinoshita, M., Honda, E., Furuta, I., & Kakehi, K. (2003). A new fungal lectin recognizing α(1-6)-linked fucose in the N-glycan. Journal of Biological Chemistry, 278(34), 32439-32447. https://doi.org/10.1074/jbc.M305181200