title: A clostridial endo-beta-galactosidase that cleaves both blood group A and B glycotopes: the first member of a new glycoside hydrolase family, GH98.
abstract: We have isolated an endo-beta-galactosidase designated E-ABase from Clostridium perfringens ATCC 10543 capable of liberating both the A trisaccharide (A-Tri; GalNAcalpha1-->3(Fucalpha1-->2)Gal) and B trisaccharide (B-Tri; Galalpha1-->3(Fucalpha1-->2)Gal) from glycoconjugates containing blood group A and B glycotopes, respectively. We have subsequently cloned the gene (eabC) that encodes E-ABase from this organism. This gene was found to be identical to the CPE0329 gene of C. perfringens strain 13, whose product was labeled as a hypothetical protein (Shimizu, T., Ohtani, K., Hirakawa, H., Ohshima, K., Yamashita, A., Shiba, T., Ogasawara, N., Hattori, M., Kuhara, S., and Hayashi, H. (2002) Proc. Natl. Acad. Sci. U. S. A. 99, 996-1001). Since the amino acid sequence of E-ABase does not bear detectable similarity to any of the 97 existing families of glycoside hydrolases, we have proposed to assign this unusual enzyme to a new family, GH98. We also expressed eabC in Escherichia coli BL21(DE3) and obtained 27 mg of fully active recombinant E-ABase from 1 liter of culture. Recombinant E-ABase not only destroyed the blood group A and B antigenicity of human type A and B erythrocytes, but also released A-Tri and B-Tri from blood group A(+)- and B(+)- containing glycoconjugates. The structures of A-Tri and B-Tri liberated from A(+) porcine gastric mucin and B(+) human ovarian cyst glycoprotein were established by NMR spectroscopy. The unique specificity of E-ABase should make it useful for studying the structure and function of blood group A- and B-containing glycoconju-gates as well as for identifying other glycosidases belonging to the new GH98 family.
authors.affiliation: Department of Biochemistry, Tulane University Health Sciences Center School of Medicine, New Orleans, Louisiana 70112, USA.
authors.forename: Kimberly M
authors.initials: KM
authors.lastname: Anderson
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ashida
authors.forename: Karol
authors.initials: K
authors.lastname: Maskos
authors.forename: Anne
authors.initials: A
authors.lastname: Dell
authors.forename: Su-Chen
authors.initials: SC
authors.lastname: Li
authors.forename: Yu-Teh
authors.initials: YT
authors.lastname: Li
mesh_terms: D000017:ABO Blood-Group System; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002469:Cell Separation; D002845:Chromatography; D002850:Chromatography, Gel; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D003016:Clostridium perfringens / Q000201:enzymology*; D003208:Concanavalin A / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D019992:Databases as Topic; D004591:Electrophoresis, Polyacrylamide Gel; D004912:Erythrocytes / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology; D005260:Female; D005434:Flow Cytometry; D006023:Glycoproteins / Q000737:chemistry; D006026:Glycoside Hydrolases / Q000737:chemistry*; D006801:Humans; D006868:Hydrolysis; D037102:Lectins / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010048:Ovarian Cysts / Q000378:metabolism; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011134:Polysaccharides / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D013552:Swine; D013997:Time Factors
chemical_list: D000017:ABO Blood-Group System; D017931:DNA Primers; D006023:Glycoproteins; D037102:Lectins; D010455:Peptides; D011134:Polysaccharides; D011994:Recombinant Proteins; C027112:fucose-binding lectin; D003208:Concanavalin A; D006026:Glycoside Hydrolases; C026093:keratan-sulfate endo-1,4-beta-galactosidase