Purification and characterization of Dolichos lablab lectin |
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Authors: | Mo H; Meah Y; Moore JG; Goldstein IJ |
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Institution: | Department of Biological Chemistry, The University of Michigan, Ann Arbor, MI 48109-0606, USA. |
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Abstract: | The mannose/glucose-binding Dolichos lablab lectin (designated DLL) has
been purified from seeds of Dolichos lablab (hyacinth bean) to
electrophoretic homogeneity by affinity chromatography on an ovalbumin-
Sepharose 4B column. The purified lectin gave a single symmetric protein
peak with an apparent molecular mass of 67 kDa on gel filtration
chromatography, and five bands ranging from 10 kDa to 22 kDa upon SDS-PAGE.
N-Terminal sequence analysis of these bands revealed subunit heterogeneity
due to posttranslational proteolytic truncation at different sites mostly
at the carboxyl terminus. The carbohydrate binding properties of the
purified lectin were investigated by three different approaches:
hemagglutination inhibition assay, quantitative precipitation inhibition
assay, and ELISA. On the basis of these studies, it is concluded that the
Dolichos lablab lectin has neither an extended carbohydrate combining site,
nor a hydrophobic binding site adjacent to it. The carbohydrate combining
site of DLL appears to most effectively accommodate a nonreducing terminal
alpha-d-mannosyl unit, and to be complementary to the C-3, C-4, and C-6
equatorial hydroxyl groups of alpha-d-mannopyranosyl and
alpha-d-glucopyranosyl residues. DLL strongly precipitates murine IgM but
not IgG, and the recent finding that this lectin interacts specifically
with NIH 3T3 fibroblasts transfected with the Flt3 tyrosine kinase receptor
and preserves human cord blood stem cells and progenitors in a quiescent
state for prolonged periods in culture, make this lectin a valuable tool in
biomedical research.
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