title: Engineering a versatile tandem repeat-type alpha2-6sialic acid-binding lectin.
abstract: Previously, we developed an alpha2-6-sialic acid (Sia)-specific lectin (SRC) starting from an R-type galactose-specific lectin C-terminal domain. However, it showed relatively low affinity because of its monovalency. Here, we engineered a tandem repeat construct (SRC2) showing substantial affinity for alpha2,6-sialylated N-glycans (in the order of 10(-6)M in K(d)), almost comparable to a natural alpha2-6Sia-specific lectin from Sambucus sieboldiana (SSA). Notably, its binding to branched N-glycans was found to be more selective than SSA. Nevertheless, SRC2 showed no apparent hemagglutinating activity, while it exerted strong erythrocyte-binding activity. This unique feature will help flow cytometry analysis, where usual lectins including SSA agglutinate cells. Some other biochemical properties investigated for SRC2, e.g., high productivity in bacteria and easy release of captured glycoproteins with lactose have demonstrated versatility of this mutant protein as a powerful tool for sialoglycomics.
authors.affiliation: Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki 305-8568, Japan.
authors.forename: Rikio
authors.initials: R
authors.lastname: Yabe
authors.forename: Yoko
authors.initials: Y
authors.lastname: Itakura
authors.forename: Sachiko
authors.initials: S
authors.lastname: Nakamura-Tsuruta
authors.forename: Jun
authors.initials: J
authors.lastname: Iwaki
authors.forename: Atsushi
authors.initials: A
authors.lastname: Kuno
authors.forename: Jun
authors.initials: J
authors.lastname: Hirabayashi
mesh_terms: D002460:Cell Line; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D037102:Lectins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019158:N-Acetylneuraminic Acid / Q000737:chemistry / Q000378:metabolism; D037121:Plant Lectins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D015202:Protein Engineering; D027923:Sambucus / Q000737:chemistry / Q000378:metabolism; D020080:Tandem Repeat Sequences / Q000235:genetics
chemical_list: D006023:Glycoproteins; D037102:Lectins; D037121:Plant Lectins; D019158:N-Acetylneuraminic Acid