Inhibitory effects and specificity of synthetic sialyldendrimers toward recombinant human cytosolic sialidase 2 (NEU2).
Human sialidase 2 (NEU2) is a cytoplasmic sialidase that degrades sialylglycoconjugates, including glycoproteins and gangliosides, via hydrolysis of terminal sialic acids to produce asialo-type molecules. Here, we first report the inhibitory effects of a series of synthetic sialyldendrimers comprising three types [Dumbbell(1)6-S-Neu5Ac(6), Fan(0)3-S-Neu5Ac(3) and Ball(0)4-S-NeuAc(4)] toward recombinant human NEU2 in vitro. Among them, Dumbbell(1)6-S-Neu5Ac(6) exhibited the most potent inhibitory activity (concentration causing 50% inhibition (IC(50)), 0.4 ∼ 0.5 mM). In addition, NeuSLac and NeuSCel carrying thiosialyltrisaccharide moieties exhibited more potent inhibitory effects than NeuSGal and NeuSGlc carrying thiosialyldisaccharides. Docking models composed of NEU2 and the thiosialyloligosaccharide suggested that the active pocket of NEU2 prefers the second galactose-β (Galβ) to the glucose-β (Glcβ) residue in the trisaccharide structure, there being a hydrogen bond between the 4-hydroxy group of the second Galβ and the side chain of the D46 residue of NEU2. The third Glcβ residues of NeuSLac and NeuSCel were also predicted to be stabilized by hydrogen bonds with the side chains of the R21, R304, D358 and Y359 residues of NEU2. NEU2 mutants (D358A and Y359A) exhibited reduced affinity for NeuSLac carrying thiosialyltrisaccharide moieties, suggesting the significant roles of D358 and Y359 residues in recognition of thiosialyltrisaccharide moieties of NeuSLac bound in the active pocket of NEU2. Thus, the present sialyldendrimers could be utilized not only as a new class of NEU2 inhibitors but also as molecular probes for evaluating the biological functions of NEU2, including the catalytic activity and mechanism as to natural substrates carrying sialyloligosaccharides.
Department of Medicinal Biotechnology, Institute for Medicinal Research, Graduate School of Pharmaceutical Sciences, The University of Tokushima, Tokushima 770-8505, Japan.
M Motiur
MM
Rahman
Satoshi
S
Kitao
Daisuke
D
Tsuji
Kaori
K
Suzuki
Jun-Ichi
J
Sakamoto
Koji
K
Matsuoka
Fumiko
F
Matsuzawa
Sei-Ichi
S
Aikawa
Kohji
K
Itoh
D000595:Amino Acid Sequence; D001665:Binding Sites; D002240:Carbohydrate Sequence; D050091:Dendrimers / Q000737:chemistry*; D004791:Enzyme Inhibitors / Q000737:chemistry* / Q000494:pharmacology; D005690:Galactose / Q000737:chemistry; D005947:Glucose / Q000737:chemistry; D006860:Hydrogen Bonding; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D019158:N-Acetylneuraminic Acid / Q000737:chemistry*; D009439:Neuraminidase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors; D013379:Substrate Specificity; D014312:Trisaccharides / Q000737:chemistry
D050091:Dendrimers; D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D014312:Trisaccharides; C403285:NEU2 protein, human; D009439:Neuraminidase; D019158:N-Acetylneuraminic Acid; D005947:Glucose; D005690:Galactose