title: Cytotoxicity and glycan-binding profile of a D-galactose-binding lectin from the eggs of a Japanese sea hare (Aplysia kurodai).
abstract: A divalent cation-independent 16 kDa D-galactose binding lectin (AKL-2) was isolated from eggs of sea hare, Aplysia kurodai. The lectin recognized D-galactose and D-galacturonic acid and had a 32 kDa dimer consisting of two disulfide-bonded 16 kDa subunits. Eighteen N-terminus amino acids were identified by Edman degradation, having unique primary structure. Lectin blotting analysis with horseradish peroxidase-conjugated lectins has shown that AKL-2 was a glycoprotein with complex type oligosaccharides with N-acetyl D-glucosamine and mannose at non-reducing terminal. Two protein bands with 38 and 36 kDa in the crude extract of sea hare eggs after purification of the lectin was isolated by AKL-2-conjugated Sepharose column and elution with 0.1 M lactose containing buffer. It suggested that the lectin binds with an endogenous ligand in the eggs. AKL-2 kept extreme stability on haemagglutination activity if it was treated at pH 3 and 70 °C for 1 h. Glycan binding profile of AKL-2 by frontal affinity chromatography technology using 15 pyridylamine labeled oligosaccharides has been appeared that the lectin uniquely recognized globotriose (Galα1-4Galβ1-4Glc; Gb3) in addition to bi-antennary complex type N-linked oligosaccharides with N-acetyllactosamine. Surface plasmon resonance analysis of AKL-2 against a neo-glycoprotein, Gb3-human serum albumin showed the k(ass) and k(diss) values are 2.4 × 10³ M⁻¹ s⁻¹ and 3.8 × 10⁻³ s⁻¹, respectively. AKL-2 appeared cytotoxicity against both Burkitt's lymphoma Raji cell and erythroleukemia K562. The activity to Raji by the lectin was preferably cancelled by the co-presence of melibiose mimicing Gb3. On the other hand, K562 was cancelled effectively by lactose than melibiose. It elucidated that AKL-2 had cytotoxic ability mediated glycans structure to cultured cells.
authors.affiliation: Department of Chemistry, Graduate School of Science, The University of Chittagong, Chittagong 4331, Bangladesh.
authors.forename: Sarkar M A
authors.initials: SM
authors.lastname: Kawsar
authors.forename: Ryo
authors.initials: R
authors.lastname: Matsumoto
authors.forename: Yuki
authors.initials: Y
authors.lastname: Fujii
authors.forename: Haruki
authors.initials: H
authors.lastname: Matsuoka
authors.forename: Naoko
authors.initials: N
authors.lastname: Masuda
authors.forename: Iwahara
authors.initials: I
authors.lastname: Chihiro
authors.forename: Hidetaro
authors.initials: H
authors.lastname: Yasumitsu
authors.forename: Robert A
authors.initials: RA
authors.lastname: Kanaly
authors.forename: Shigeki
authors.initials: S
authors.lastname: Sugawara
authors.forename: Masahiro
authors.initials: M
authors.lastname: Hosono
authors.forename: Kazuo
authors.initials: K
authors.lastname: Nitta
authors.forename: Naoto
authors.initials: N
authors.lastname: Ishizaki
authors.forename: Chikaku
authors.initials: C
authors.lastname: Dogasaki
authors.forename: Jiharu
authors.initials: J
authors.lastname: Hamako
authors.forename: Taei
authors.initials: T
authors.lastname: Matsui
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Ozeki
mesh_terms: D000818:Animals; D001048:Aplysia / Q000737:chemistry / Q000378:metabolism*; D045744:Cell Line, Tumor; D037161:Galectins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity*; D006801:Humans; D007564:Japan; D007700:Kinetics; D008970:Molecular Weight; D010063:Ovum / Q000737:chemistry / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D011485:Protein Binding
chemical_list: D037161:Galectins; D011134:Polysaccharides