The amino acids involved in the distinct carbohydrate specificities between macrophage galactose-type C-type lectins 1 and 2 (CD301a and b) of mice.
Binding specificities of mouse macrophage galactose-type C-type lectin 1 (MGL1/CD301a) and 2 (MGL2/CD301b) toward various oligosaccharides were compared by frontal affinity chromatography. MGL1 preferentially bound oligosaccharides containing Lewis(X) (Le(X)) trisaccharides among 111 oligosaccharides tested, whereas MGL2 preferentially bound globoside Gb4. The important amino acids for the preferential bindings were investigated by pair-wise site-directed mutagenesis at positions 61, 89, 97, 100, 110-113, 115, 124, and 125 in the soluble recombinant carbohydrate recognition domains (CRD) prepared in Escherichia coli and purified with galactose-Sepharose. Mutations of Val, Ala, Thr, and Phe at positions 61, 89, 111 and 125 on MGL1 CRD caused reductions in Le(X) binding. Mutations of MGL2 CRD at Leu, Arg, Arg, and Tyr at positions 61, 89, 115 and 125 were implicated in the preference for beta-GalNAc. Le(X) binding was observed with MGL2 mutants of Arg89Ala and Arg89Ala/Ser111Thr. MGL1 mutants of Ala89Arg and Ala89Arg/Pro115Arg showed beta-GalNAc bindings. Molecular modeling illustrated potential direct molecular interactions of Leu61, Arg89, and His109 in MGL2 CRD with GalNAc.
Oo-Puthinan
Sarawut
S
Laboratory of Cancer Biology and Molecular Immunology, The University of Tokyo, Tokyo 113-0033, Japan.
Maenuma
Keisuke
K
Sakakura
Masayoshi
M
Denda-Nagai
Kaori
K
Tsuiji
Makoto
M
Shimada
Ichio
I
Nakamura-Tsuruta
Sachiko
S
Hirabayashi
Jun
J
Bovin
Nicolai V
NV
Irimura
Tatsuro
T
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000596:Amino Acids / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D000818:Animals; D001212:Asialoglycoproteins / Q000235:genetics / Q000276:immunology*; D002241:Carbohydrates / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D037181:Lectins, C-Type / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D009844:Oligosaccharides / Q000737:chemistry / Q000276:immunology*
D000596:Amino Acids; D001212:Asialoglycoproteins; D002241:Carbohydrates; C064720:Clec10a protein, mouse; D037181:Lectins, C-Type; C464940:MGL2 protein, mouse; D008565:Membrane Proteins; D009844:Oligosaccharides