VIPL has sugar-binding activity specific for high-mannose-type N-glycans, and glucosylation of the α1,2 mannotriosyl branch blocks its binding
2007; Oxford University Press; Volume: 17; Issue: 10 Linguagem: Inglês
10.1093/glycob/cwm074
ISSN1460-2423
AutoresDaisuke Yamaguchi, Norihito Kawasaki, Ichiro Matsuo, Kiichiro Totani, Hideto Tozawa, Naoki Matsumoto, Yukishige Ito, Kazuo Yamamoto,
Tópico(s)Galectins and Cancer Biology
ResumoVIP36-like protein (VIPL) was identified as an endoplasmic reticulum (ER) resident protein with homology to VIP36, a cargo receptor involved in the transport of glycoproteins within cells. Although VIPL is structurally similar to VIP36, VIPL is thought not to be a lectin, because its sugar-binding activity has not been detected in several experiments. Here, recombinant soluble VIPL proteins (sVIPL) were expressed in Escherichia coli, biotinylated with biotin ligase and oligomerized with R-phycoerythrin (PE)-labeled streptavidin (SA). As measured with flow cytometry, PE-labeled sVIPL–SA bound to deoxymannojirimycin (DMJ)- or kifunensine (KIF)- but not to swainsonine (SW)-treated HeLaS3 cells in the presence of calcium. A surface plasmon resonance analysis showed that the avidity of sVIPL was enhanced after it formed a complex with SA. The binding of PE-labeled sVIPL–SA was abrogated by endo β-N-acetylglucosaminidase H treatment of the DMJ- or KIF-treated cells. Competition with several high-mannose-type N-glycans inhibited VIPL binding, and indicated that VIPL recognizes the Manα1–2Manα1–2Man sequence. Glucosylation of the outer mannose residue of this portion decreased the binding. Although the biochemical characteristics of VIPL are similar to those of VIP36, the sugar-binding activity of VIPL was stronger at neutral pH, corresponding to the pH in the lumen of the ER, than under acidic conditions.
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