Evaluation of Receptor Binding Specificity ofEscherichia coliK88 (F4) Fimbrial Adhesin Variants Using Porcine Serum Transferrin and Glycosphingolipids as Model Receptors

Abstract

Diarrheal disease caused by enterotoxigenic Escherichia coli expressing the K88 (F4) fimbrial adhesin (K88 ETEC) is a significant source of mortality and morbidity among newborn and weaned piglets. K88 fimbrial adhesins are filamentous surface appendages whose lectin (carbohydrate-binding) activity allows K88 ETEC to attach to specific glycoconjugates (receptors) on porcine intestinal epithelial cells. There are three variants of K88 adhesin (K88ab, K88ac, and K88ad), which possess different, yet related, carbohydrate-binding specificities. We used porcine serum transferrin (pSTf) and purified glycosphingolipids (GSL) to begin to define the minimal recognition sequence for K88 adhesin variants. We found that K88ab adhesin binds with high affinity to pSTf (dissociation constant, 75 M), while neither K88ac nor K88ad adhesin recognizes pSTf. Degradation of the N-glycan on pSTf by extensive metaperiodate treatment abolished its interaction with the K88ab adhesin, indicating that the K88ab adhesin binds to the single N-glycan found on pSTf. Using exoglycosidase digestion of the pSTf glycan, we demonstrated that K88ab adhesin recognizes N-acetylglucosamine (GlcNAc) residues in the core of the N-glycan on pSTf. All three K88 variants were found to bind preferentially to GSL containing a ␤-linked N-acetylhexosamine (HexNAc), either GlcNAc or N-acetylgalactosamine, in the terminal position or, alternatively, in the penultimate position with galactose in the terminal position. Considering the results from pSTf and GSL binding studies together, we propose that the minimal recognition sequence for the K88 adhesin variants contains a ␤-linked HexNAc. In addition, the presence of a terminal galactose ␤-linked to this HexNAc residue enhances K88 adhesin binding.Diarrheal disease caused by enterotoxigenic Escherichia coli expressing the K88 (F4) fimbrial adhesin (K88 ETEC) is a significant source of mortality and morbidity among newborn and weaned piglets (9,25,38,41,59). K88 fimbrial adhesins are filamentous surface appendages whose lectin (carbohydratebinding) activity allows K88 ETEC to attach to specific glycoconjugates (receptors) on porcine intestinal epithelial cells (28). Attachment of bacteria to their receptors on intestinal epithelial cells is an essential step in the colonization of the small intestine by ETEC.Three serologically distinguishable variants of K88 adhesin (K88ab, K88ac, and K88ad) have been identified (21, 40). Each variant consists of a conserved antigenic region shared by all three variants, designated a, and variant-specific antigenic regions, designated b, c, and d for K88ab, K88ac, and K88ad, respectively (13, 29). The antigenic differences among the three K88 variants can be ascribed exclusively to a small number of nucleotide changes in the major fimbrial subunit gene, resulting in the amino acid substitutions that distinguish the three K88 variants. Several researchers have investigated the molecular interaction of K88 fimbrial adhesin variants with erythrocytes, intestinal mucus, and intestina...

Biorecognition of Escherichia coli K88 adhesin for glycated porcine albumin

“…Furthermore, Payne et al [47] and Grange et al [5] determined that ␤-linked galactose is an important component in the recognition of intestinal mucus K88 receptors. K88 adhesin also interacts with lactosylated (Gal ␤ 1-4 GlcNAc) N-oligosaccharides present in serum and intestinal glycoproteins [29,48,49].…”

Enterotoxigenic Escherichia coli strains are highly prevalent in Ugandan piggeries but disease outbreaks are masked by antibiotic prophylaxis

“…F4 fimbriae consist of the major adhesive subunit FaeG and the minor subunits FaeF, FaeH, FaeC and FaeI ( Van den Broeck et al 2000). The three variants of F4 fimbriae (ab, ac and ad) vary in FaeG sequence, and each shows a related but distinct binding profile to the glycoconjugate receptors on enterocytes (Grange 2002). After colonization, ETEC produce heat-labile enterotoxin (LT), heat-stable enterotoxin a and b (STa and STb) in different combinations (Fairbrother et al 2005).…”

Structural and Functional Insight into the Carbohydrate Receptor Binding of F4 Fimbriae-producing Enterotoxigenic Escherichia coli

“…The important role for galactose and/or N-acetyl galactosamine residue in the F4 receptor structure was highlighted in different studies; most often these residues were present at the non-reducing end in ␤-linkage (24, 28, 30 -36). In other studies, the interaction of all F4 variants with different glycosphingolipids was demonstrated, such as lactosylceramide, gangliotriaosylceramide, gangliotetraosylceramide, globotriaosylceramide, lactotetraosylceramide, and lactotetraosylceramide (24,30,32,37). A more recent study investigated the glycosphingolipid recognition by the different F4 variants and found that the F4 ab and F4 ac variants showed more similarities in their glycosphingolipid recognition patterns when compared with the F4 ad variant (30).…”