Selective removal of heparan sulfate chains from proteoheparan sulfate with a commercial preparation of heparitinase

Kato, Masato; Oike, Y; Suzuki, Sakaru; Kimata, Koji

doi:10.1016/0003-2697(85)90255-6

Cited by 58 publications

(20 citation statements)

References 22 publications

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“…Although the tight binding of H A to protein cores of proteoglycans has been shown in hyaline cartilage [21], it does not seem likely under the present dissociative elution condition, such as 8 M urea with 0.5% Triton X-100 [15,201. Recently,Kato et al reported that the commercial preparation of heparitinase has a significant proteolytic activity [22]. However, under the present digestive condition, the enzyme does not seem to have any significant proteolytic activity, since both peak I and I1 fractions were not degraded at all by the enzyme.…”

Section: Discussionmentioning

confidence: 97%

Secretions from Primary Hamster Tracheal Surface Epithelial Cells in Culture: Mucin-Like Glycoproteins, Proteoglycans, and Lipids

Opaskar-Hincman

1989

Experimental Lung Research

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Surface epithelial cells dissociated from hamster tracheas and grown on a thick collagen gel in the presence of 5% fetal bovine serum become highly enriched with secretory cells at confluence. In the present communication, we have analyzed secretory products from this primary hamster tracheal surface epithelial (HTSE) cell culture. The secreted glycoconjugates included high-molecular-weight mucin-like glycoproteins (HMW MLGP) and proteoglycans that comprised 22% and 5% of the total [3H]glycoconjugates secreted when [3H]glucosamine was added as a metabolic precursor. Among the proteoglycans were hyaluronic acids (53%), heparan sulfate proteoglycans (29%), and chrondroitin sulfate proteoglycans (18%). Chondroitin sulfates were mostly 4-sulfated. On the other hand, the secreted lipids included cholesterol, phospholipids, and glycolipids, and most of them were associated with HMW MLGP.

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Section: Discussionmentioning

confidence: 97%

Secretions from Primary Hamster Tracheal Surface Epithelial Cells in Culture: Mucin-Like Glycoproteins, Proteoglycans, and Lipids

Opaskar-Hincman

1989

Experimental Lung Research

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“…The glycoproteins were treated with 0.2 units/ml of heparitinase I or chondroitinase ABC (Seikagaku Kogyo Co., Tokyo, Japan) in 30 mM Tris-acetate buffer, pH 7.5, containing 10 mM EDTA, 10 mM NEM, 1 mM PMSF and pepstatin-A (0.25 mg/ml) at 37°C for 1 h according to Kato et al (1985). Sialidase (Nacalai Tesque, Kyoto, Japan) treatment (0.06 units/ml) was performed in 30 mM sodium acetate buffer, pH 5.0, containing 5 mM EDTA, 5 mM NEM, 1 mM PMSF and pepstatin-A (0.07 mg/ml) at 37°C for 20 h. N-Glycosidase-F (Takara Shuzo, Ohtsu, Japan) treatment (0.04 units/ml) was performed at 37°C for 20 h according to manufacturer's instructions.…”

Section: Enzymatic Treatments With Glycosidases and Proteasesmentioning

confidence: 99%

Identification of the Core Protein Carrying the Tn Antigen in Mouse Brain: Specific Expression on Syndecan-3.

Akita

Fushiki

Fujimoto

et al. 2001

Cell Struct. Funct.

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ABSTRACT. We isolated glycoproteins carrying the Tn antigen, which was expressed spatiotemporally in the developing mouse brain. The Tn antigen was expressed on two molecular species with a molecular weight from 200 to 350 kDa and 110 to 160 kDa, as judged on SDS-PAGE. Although the two glycoproteins showed different susceptibilities to heparitinase I and solubilities in a salt solution, after treatment with V8 protease they showed the same mobility corresponding to a molecular weight of 90 kDa on SDS-PAGE, suggesting that these two molecules shared a common core protein. Partial N-terminal sequences of the glycoproteins were determined, i.e. AQRXRNENFERPV and ALAAPXAPAMLP, which were identified as the sequences of the N-terminal and central portions of syndecan-3, respectively. Both glycoproteins were reactive to anti-mouse syndecan-3 antibody. These results suggest that one is a soluble syndecan-3 cleaved between mucin-like domain and transmembrane domain, and the other is a membrane-bound syndecan-3 lacking N-terminal glycosaminoglycan attachment sites, and that both glycoproteins have a mucin-like domain characteristic of syndecan-3, in which the Tn antigen may be expressed.

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“…However, we have had no problems in the electrophoretic transfer of PC12 cell proteoglycans to nitrocellulose, and the monoclonal antibodies reported to recognize the HSPG protein core have not required prior removal ofheparan sulfate chains for reactivity (7). Since flavobacterial heparan sulfatedegrading enzymes are known to contain contaminating protease activity, and since the enzyme digestions were apparently performed in the absence of protease inhibitors (11), an alternative explanation for the findings of Schubert et al not addressed in their report is that the 65-kD immunoreactive band (whose sequence was not determined) may have been generated from a larger protein as a result of proteolysis.…”

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confidence: 89%

Relation of the Amyloid β Protein Precursor to Heparan Sulfate Proteoglycans

Gowda

Frangione

Ghiso

et al. 1989

Science

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any case their presence does not detract from the differences between the leukemic and remission samples of this patient. Miller et al. and Donti et al. suggest that the conclusions of our paper imply an understanding of the pathogenesis of APL or a direct association between rearranged MPO and critical DNA sequences involved in the t(15;17); this is not the case. Although questioned, it should be noted that only "one explanation for the presence of these novel bands in two of the M3 (APL) patients examined is that the breakpoint on chromosome 17 of the t(15;17) occurs within the MPO sequences. Alternatively, M3-associated rearrangements of the MPO gene such as submicroscopic deletions or inversions that are unrelated to the translocation breakpoint may have occurred" (1).We were not aware ofthe publications by Chang et al. in Leukemia (2) In the course of our previous studies of proteoglycans and other glycoconjugates in this neuronal cell line (2-4), we obtained data that raise several questions concerning this conclusion.1) Schubert et al. state that they purified the secreted form of a neuronal HSPG from PC12 cells and that they used this purified HSPG to obtain tryptic peptides that were fractionated by high-performance liquid chromatography. A major peptide labeled with 35SO4 (designated peptide 51) had an NH2-terminal amino acid sequence that was almost identical to the deduced sequence for residues 18 through 44 of human ABPP. The authors conclude that this peptide must be derived from an HSPG core protein, as HSPGs are the only proteoglycans produced by PC12 cells, and that all tryptic peptides labeled with 35s04 should therefore be derived from this proteoglycan.However, we previously reported (2) that chondroitin sulfate accounts for approximately 80% of the glycosaminoglycans secreted (in the form of proteoglycans) by the original line of PC12 cells (5), and we have recently found (4, 6) that chondroitin sulfate proteoglycans represent a smaller but still highly significant proportion (approximately 35%) ofthe proteoglycans secreted by the PC12 cells studied in two other laboratories (1, 7). The paper (7) cited as evidence that only one proteoglycan has been detected in PC12 cells reports an investigation specifically of HSPGs identified with monoclonal antibodies and does not address the question of whether other proteoglycans may also be present. Moreover, the reference cited by Schubert et al. in support of the statement that HSPGs are the major class of proteoglycans in nervous tissue clearly demonstrates that chondroitin sulfate, and not heparan sulfate, is the predominant sulfated glycosaminoglycan at all ages in brain (8).Because we have found [on the basis of gel filtration and SDS-PAGE (polyacrylamide gel electrophoresis)] that the molecular size of the HSPG secreted by PC12 cells (110 to 135 kD) is the same as that recently reported for the ABPP (9), we do not think it is surprising that these would elute together on Sepharose CL-4B. Therefore, in the absence of electrophoretic and fluo...

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Selective removal of heparan sulfate chains from proteoheparan sulfate with a commercial preparation of heparitinase

Cited by 58 publications

References 22 publications

Secretions from Primary Hamster Tracheal Surface Epithelial Cells in Culture: Mucin-Like Glycoproteins, Proteoglycans, and Lipids

Secretions from Primary Hamster Tracheal Surface Epithelial Cells in Culture: Mucin-Like Glycoproteins, Proteoglycans, and Lipids

Identification of the Core Protein Carrying the Tn Antigen in Mouse Brain: Specific Expression on Syndecan-3.

Relation of the Amyloid β Protein Precursor to Heparan Sulfate Proteoglycans

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