The antigenic determinants recognized by three monoclonal antibodies to keratan sulphate involve sulphated hepta- or larger oligosaccharides of the poly(N-acetyllactosamine) series

Mehmet, Huseyin; Scudder, Peter; Tang, Ping; Hounsell, Elizabeth F.; Caterson, Bruce; Feizi, Ten

doi:10.1111/j.1432-1033.1986.tb09680.x

Cited by 226 publications

(134 citation statements)

References 33 publications

(21 reference statements)

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“…Previously, we found comparatively low levels of sulfated KS in normal mouse cornea based on immunoreactivity with 5D4 (22), a mAb that recognizes high sulfated sequences of residues (minimally penta-sulfated) on poly N-acetyllactosamine disaccharides of KS (23,24). In the current study these KS epitopes were detected in extracts of WT and heterozygous corneas, but not in corneal extracts from homozygous Chst5-null mice (Fig.…”

Section: Resultssupporting

confidence: 50%

Matrix morphogenesis in cornea is mediated by the modification of keratan sulfate by GlcNAc 6- O -sulfotransferase

Hayashida

Akama

Beecher

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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Matrix assembly and homeostasis in collagen-rich tissues are mediated by interactions with proteoglycans (PGs) substituted with sulfated glycosaminoglycans (GAGs). The major GAG in cornea is keratan sulfate (KS), which is N-linked to one of three PG core proteins. To ascertain the importance of the carbohydrate chain sulfation step in KS functionality, we generated a strain of mice with a targeted gene deletion in Chst5, which encodes an N-acetylglucosamine-6-O-sulfotransferase that is integral to the sulfation of KS chains. Corneas of homozygous mutants were significantly thinner than those of WT or heterozygous mice. They lacked high-sulfated KS, but contained the core protein of the major corneal KSPG, lumican. Histochemically stained KSPGs coassociated with fibrillar collagen in WT corneas, but were not identified in the Chst5-null tissue. Conversely, abnormally large chondroitin sulfate͞dermatan sulfate PG complexes were abundant throughout the Chst5-deficient cornea, indicating an alteration of controlled PG production in the mutant cornea. The corneal stroma of the Chst5-null mouse exhibited widespread structural alterations in collagen fibrillar architecture, including decreased interfibrillar spacing and a more spatially disorganized collagen array. The enzymatic sulfation of KS GAG chains is thus identified as a key requirement for PG biosynthesis and collagen matrix organization.collagen ͉ glycosaminoglycans ͉ proteoglycans G lycosaminoglycans (GAGs) substituted on proteoglycans (PGs) are influential in defining collagen fibrillar architecture in a wide range of connective tissue matrices. Keratan sulfate (KS) is an important constituent of several collagen-rich tissues and is the major GAG in cornea where it is N-linked to asparagine residues in one of three PG core proteins: lumican (1), keratocan (2), and mimecan͞osteoglycin (3). Human corneal GlcNAc 6-O-sulfotransferase (also known as human GlcNAc6ST-5 and GST4␤) is the responsible enzyme for the synthesis of high-sulfated KS via the transfer of sulfate onto the GlcNAc 6-O position of the KS backbone (4).Fairly compelling evidence exists for a regulatory role for KSPGs in the maintenance of corneal matrix structure in a number of species. The avian cornea in ovo, for example, synthesizes an unsulfated form of KS midway through development when it is structurally disorganized and transmits relatively little light, but switches to produce a sulfated KS GAG as it becomes transparent and attains a more well ordered collagen fibrillar ultrastructure (5, 6). KS sulfation patterns are also altered in opaque, structurally disorganized corneal scar tissue in rabbits (7,8) and in cloudy human corneas with the inherited disease, macular corneal dystrophy (9), which is caused by mutations in CHST6, a gene encoding human corneal GlcNAc 6-O-sulfotransferase (10).Hybrid type I͞V collagen fibrils are the cornea's main nonspecular light-scattering elements and are formed into wide, interweaving belts or lamellae that lie approximately in the tissue plane (11). Wit...

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Section: Resultssupporting

confidence: 50%

Matrix morphogenesis in cornea is mediated by the modification of keratan sulfate by GlcNAc 6- O -sulfotransferase

Hayashida

Akama

Beecher

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…After reduction and alkylation, analysis using dot-blots (see below) showed that the G1 preparation reacted with MAb 12/21/1-C-6 (to a common peptide epitope in both the G1 and G2 domains of the core protein [21]), 9/30/8-A-4 (to a peptide epitope present in link protein [17]), and 1/20/5-D-4 (to oligosaccharides from keratan sulfate consisting of several repeats of the disulfated disaccharide GlcNAc[6-sulfate] pl-3 Gal[6-sulfate] pl-4 [23]). This fraction did not react with MAb 5/6/3-B-3 (to terminal unsaturated chondroitin-6-sulfate disaccharides [17]), which indicated that the G1 preparation contained mainly G1, with some link protein present.…”

Section: Methodsmentioning

confidence: 99%

Presence of antibodies to native G1 domain of aggrecan core protein in synovial fluids from patients with various joint diseases

Karopoulos

Rowley

Ilic

et al. 1996

Arthritis & Rheumatism

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Objective. To investigate the occurrence of IgG antibodies to aggrecan in synovial fluids (SF) from patients with arthritis and various articular diseases, and to determine the nature of epitopes present within aggrecan that react with these antibodies. Methods. SF samples were reacted with native aggrecan, reduced and alkylated aggrecan, chondroitin sulfate, and keratan sulfate, using dot‐blots and a novel enzyme‐linked immunosorbent assay (ELISA). The nature of the epitopes present on aggrecan was elucidated using Western blots and a competitive inhibition ELISA. Results. IgG antibodies to aggrecan were found in >50% of the SF samples tested. No IgG antibody reactivity was observed in serum from the same patients. The antibodies appeared to react predominantly with native aggrecan, and there was no disease specificity. It was shown that the epitopes to these antibodies were located within the N‐terminal region of the core protein. Conclusion. This study demonstrates the frequent occurrence of IgG antibodies to aggrecan in human SF. The major epitope is located in the G1 domain of the aggrecan core protein. These IgG antibodies appear to be produced locally within the synovial cavity, probably in response to various articular diseases, resulting in the loss of native aggrecan from articular cartilage.

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“…Under normal culture conditions, SV40-transformed human corneal epithelial cells do not produce detectable levels of highly sulfated KS GAG, which is recognized by the 5D4 monoclonal antibody (28,29). However, the corneal cells began producing 5D4-positive, highly sulfated KS carbohydrate when CGn6ST and KSG6ST were overexpressed (Fig.…”

Section: Tablementioning

confidence: 99%

Enzymes Responsible for Synthesis of Corneal Keratan Sulfate Glycosaminoglycans

Kitayama

Hayashida

Nishida

et al. 2007

Journal of Biological Chemistry

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Keratan sulfate glycosaminoglycans are among the most abundant carbohydrate components of the cornea and are suggested to play an important role in maintaining corneal extracellular matrix structure. Keratan sulfate carbohydrate chains consist of repeating N-acetyllactosamine disaccharides with sulfation on the 6-O positions of N-acetylglucosamine and galactose. Despite its importance for corneal function, the biosynthetic pathway of the carbohydrate chain and particularly the elongation steps are poorly understood. Here we analyzed enzymatic activity of two glycosyltransferases, ␤1,3-N-acetylglucosaminyltansferase-7 (␤3GnT7) and ␤1,4-galactosyltransferase-4 (␤4GalT4), in the production of keratan sulfate carbohydrate in vitro. These glycosyltransferases produced only short, elongated carbohydrates when they were reacted with substrate in the absence of a carbohydrate sulfotransferase; however, they produced extended GlcNAc-sulfated poly-Nacetyllactosamine structures with more than four repeats of the GlcNAc-sulfated N-acetyllactosamine unit in the presence of corneal N-acetylglucosamine 6-O sulfotransferase (CGn6ST). Moreover, we detected production of highly sulfated keratan sulfate by a two-step reaction in vitro with a mixture of ␤3GnT7/ ␤4GalT4/CGn6ST followed by keratan sulfate galactose 6-O sulfotransferase treatment. We also observed that production of highly sulfated keratan sulfate in cultured human corneal epithelial cells was dramatically reduced when expression of ␤3GnT7 or ␤4GalT4 was suppressed by small interfering RNAs, indicating that these glycosyltransferases are responsible for elongation of the keratan sulfate carbohydrate backbone.

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The antigenic determinants recognized by three monoclonal antibodies to keratan sulphate involve sulphated hepta- or larger oligosaccharides of the poly(N-acetyllactosamine) series

Cited by 226 publications

References 33 publications

Matrix morphogenesis in cornea is mediated by the modification of keratan sulfate by GlcNAc 6- O -sulfotransferase

Matrix morphogenesis in cornea is mediated by the modification of keratan sulfate by GlcNAc 6- O -sulfotransferase

Presence of antibodies to native G1 domain of aggrecan core protein in synovial fluids from patients with various joint diseases

Enzymes Responsible for Synthesis of Corneal Keratan Sulfate Glycosaminoglycans

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