Isolation of two blood type A and N specific isolectins from Moluccella laevis seeds

FEBS Letters

Lis, Halina¹  Sharon, Nathan¹  Adar, Rivka¹  Latter, Hedva¹ 

[1] Department of Biophysics, The Weizmann Institute of Science, Rehovot 76100, Israel

关键词: Lectin;    Subunit;    Disulfide bridge;    Blood type specificity;    Sugar specificity;    HPLC;    high-performance liquid chromatography;    PAGE;    polyacrylamide gel electrophoresis;    PBS;    phosphate-buffered saline;    pH 7.4;   

DOI  :  10.1016/0014-5793(88)81382-6

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Extraction of Moluccella laevis seed flour with phosphate-buffered saline, followed by ammonium sulfate (40%) precipitation and affinity chromatography of the dialysed precipitate on a column of galactose-bound Sepharose, afforded a single protein peak with high agglutinating activity for human A^MM and O^NN erythrocytes and poor activity for B^MM and O^MM cells. Gel filtration of the affinity-purified lectin on a column of Superose 12 gave two peaks that differed markedly in their activities for A^MM and O^NN erythrocytes. Whereas the ratio of A/N activities in the slow-moving peak (isolectin I) was 0.25-0.5, that of the fast-moving peak (isolectin II) was 2–3. Molecular mass estimation by gel filtration on Sephadex G-150 gave a value of 130 kDa for isolectin I and 240 kDa for isolectin II. SDS-polyacrylamide gel electrophoresis of isolectin I without or with mercaptoethanol revealed that it contains a subunit of 67 kDa, consisting of two chains (46 and 28 kDa, after reduction) held together by SS bonds, and two noncovalently linked subunits of 42 and 26 kDa. The same subunits are also present in isolectin II, where the 67 kDa seems to be predominant. The M. laevis isolectins are unique in that they are specific for determinants of two different blood group systems, and also because they contain both covalently and noncovalently linked subunits. The pattern of inhibition by different sugars of the two blood type activities of the isolectins, assayed with either A^MM or O^NN erythrocytes, was similar. Of the monosaccharides tested, N-acetylgalactosamine was the best inhibitor, being 300–600 times more active than galactose. Methyl α-galactoside was a better inhibitor than the corresponding β-galactoside. The sugar specificity fits well with the specifity of the isolectins for blood type A erythrocytes, but does not account for their type N specificity.

【 授权许可】

Unknown