RRC ID 28630
著者 Nishi N, Abe A, Iwaki J, Yoshida H, Itoh A, Shoji H, Kamitori S, Hirabayashi J, Nakamura T.
タイトル Functional and structural bases of a cysteine-less mutant as a long-lasting substitute for galectin-1.
ジャーナル Glycobiology
Abstract Galectin-1 (Gal-1), a member of the beta-galactoside-binding animal lectin family, has a wide range of biological activities, which makes it an attractive target for medical applications. Unlike other galectins, Gal-1 is susceptible to oxidation at cysteine residues, which is troublesome for in vitro/vivo studies. To overcome this problem, we prepared a cysteine-less mutant of Gal-1 (CSGal-1) by substituting all cysteine residues with serine residues. In the case of wild-type Gal-1, the formation of covalent dimers/oligomers was evident after 10 days of storage in the absence of a reducing agent with a concomitant decrease in hemagglutination activity, while CSGal-1 did not form multimers and retained full hemagglutination activity after 400 days of storage. Frontal affinity chromatography showed that the sugar-binding specificity and affinity of Gal-1 for model glycans were barely affected by the mutagenesis. Gal-1 is known to induce cell signaling leading to an increase in the intracytoplasmic calcium concentration and to cell death. CSGal-1 is also capable of inducing calcium flux and growth inhibition in Jurkat cells, which are comparable to or more potent than those induced by Gal-1. The X-ray structure of the CSGal-1/lactose complex has been determined. The structure of CSGal-1 is almost identical to that of wild-type human Gal-1, showing that the amino acid substitutions do not affect the overall structure or carbohydrate-binding site structure of the protein. These results indicate that CSGal-1 can serve as a stable substitute for Gal-1.
巻・号 18(12)
ページ 1065-73
公開日 2008-12-1
DOI 10.1093/glycob/cwn089
PII cwn089
PMID 18796645
MeSH Amino Acid Substitution Binding Sites Cell Proliferation Crystallography, X-Ray Cysteine / genetics* Cysteine / metabolism Galectin 1 / chemistry* Galectin 1 / genetics* Galectin 1 / metabolism Humans Jurkat Cells Membrane Glycoproteins / metabolism Models, Molecular Mutagenesis, Site-Directed Mutation* Oxidation-Reduction Protein Conformation Structure-Activity Relationship
IF 4.06
引用数 51
WOS 分野 BIOCHEMISTRY & MOLECULAR BIOLOGY
リソース情報
遺伝子材料 pGEX-CSG1-2a (RDB08448) pET-CSG1-1a (RDB08449)