10E4 Antigen of Scrapie Lesions Contains an Unusual Nonsulfated Heparan Motif

Abstract

The carbohydrate antigen on heparan sulfate recognized by monoclonal antibody 10E4 is uniquely codistributed with the abnormal prion protein, PrP Sc , even in the earliest detectable brain lesions of scrapie-infected mice. Determining the chemical structure of 10E4 antigen is, therefore, an important aspect of structure elucidation of scrapie lesions, and a prerequisite for designing experiments to understand its role in scrapie pathogenesis. Toward this aim, we have examined preparations of heparan sulfate, with differing sulfate contents, for binding by 10E4 antibody. The highest antigenicity was observed in a preparation (HS-1) with the lowest sulfate content. HS-1 was partially depolymerized with heparin lyase III, and oligosaccharide fragments examined for 10E4 antigen expression by the neoglycolipid technology. An antigen-positive and two antigen-negative tetrasaccharides were isolated and examined by electrospray mass spectrometry. The antigen-positive tetrasaccharide sequence on heparan sulfate was thus deduced to contain a unique unsulfated motif that includes an N-unsubstituted glucosamine in the sequence, UA-GlcN-UA-GlcNAc. Antibody binding experiments with neoglycolipids prepared from a series of heparin/heparan sulfate disaccharides, and the trisaccharide derived from the antigen-positive tetrasaccharide after removal of the terminal hexuronic acid, show that both the penultimate glucosamine and the outer nonsulfated hexuronic acid are important for 10E4 antigenicity.

Heparan Sulfate and Transglutaminase Activity Are Required for the Formation of Covalently Cross-linked Hedgehog Oligomers

“…In these scenarios TGs have been implicated in the abnormal accumulation of insoluble and protease-resistant proteinaceous aggregates (14). In addition, HS is co-distributed with the abnormal prion protein, PrP(Sc), even in very early disease stages in scrapie-infected mice (41). This raises the possibility that HS may also serve as a scaffold for protein aggregation in these pathophysiological conditions.…”

Immunohistochemical expression of heparan sulfate correlates with stromal cell proliferation in breast phyllodes tumors

“…10E4 antibody is a widely-used antibody for the detection and localization of heparan sulfate in tissues. [26][27][28] Leteux et al 29 reported that 10E4 recognized a heparan sulfate tetrasaccharide in which an N-unsubstituted GlcN residue was required for the binding to 10E4. More recently, it was shown that a mixed N-sulfated/Nacetylated epitope was necessary for 10E4 binding.…”

Nitric Oxide-dependent Processing of Heparan Sulfate in Recycling S-Nitrosylated Glypican-1 Takes Place in Caveolin-1-containing Endosomes

“…Two HS-specific monoclonal antibodies were tested, one of which (JM-403) (29) is directed against an epitope containing GlcNH 3 ϩ . The HS epitope of the other monoclonal (10E4) has been proposed to comprise the sequence HexUA-GlcNH 3 ϩ -HexUA-GlcNAc (32). N-Acetylation of free amino groups blocked the reactivity of JM-403 (Fig.…”