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The D-domain of fibrin : structural and functional studies
The D-domain of fibrin (ogen) was separated from the parent molecule by plasmin digestion in the presence of calcium and isolated by means of DEAE-anion exchange chromatography followed by gel-filtration in buffer containing 4 M urea. Fluorescent-D-dimer (f-D-dimer) was isolated from a plasmic diges...
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| Format: | Thesis |
| Language: | English |
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Division of Chemical Pathology
2018
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| _version_ | 1867613434545176576 |
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| access_status_str | Open Access |
| author | Purves, Maud |
| author2 | Purvis, Langley R |
| author_browse | Purves, Maud Purvis, Langley R |
| author_facet | Purvis, Langley R Purves, Maud |
| author_sort | Purves, Maud |
| collection | Thesis |
| description | The D-domain of fibrin (ogen) was separated from the parent molecule by plasmin digestion in the presence of calcium and isolated by means of DEAE-anion exchange chromatography followed by gel-filtration in buffer containing 4 M urea. Fluorescent-D-dimer (f-D-dimer) was isolated from a plasmic digest of fibrin clotted in the presence of 2.45 mM dansyl cadaverine, a fluorescent lysine analogue. Fluorescent-D-monomer was a by-product of f-D-dimer purification, the yield being determined by the concentration of dansyl cadaverine. At 2.45 mM f-D-monomer was always present in the digest. The f-D-monomer is probably formed directly and not as a result of degradation of f-D-dimer. The molecule elutes in the fibrinogen-derived-D- monomer position on gel-filtration. A protease was isolated and partially purified from venom of the puffadder (Bitis arietans). Puffadder venom protease is characterized by its ability to cleave D-dimer into symmetrical D-monomers, smaller than plasmin-derived D-monomers from fibrinogen. This characteristic was used to detect the puffadder venom protease activity with fluorescent-D-dimer being used as the substrate. Fractions obtained were assayed for D-dimer cleavage activity and the samples analyzed by means of SDS-PAGE on 4-20% gradient gels under reducing (βME) and non-reducing conditions. The fluorescent bands were located under U.V light and photographed prior to staining with Coomassie Blue. Several methods for the purification of the protease were investigated. |
| format | Thesis |
| id | oai:open.uct.ac.za:11427/27202 |
| institution | University of Cape Town (South Africa) |
| language | eng |
| last_indexed | 2026-06-10T12:36:05.501Z |
| license_str | Not specified — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from UCTD — University of Cape Town Open Access Repository |
| publishDate | 2018 |
| publishDateRange | 2018 |
| publishDateSort | 2018 |
| publisher | Division of Chemical Pathology |
| publisherStr | Division of Chemical Pathology |
| record_format | dspace |
| source_str | UCTD — University of Cape Town Open Access Repository |
| spelling | oai:open.uct.ac.za:11427/27202 The D-domain of fibrin : structural and functional studies Purves, Maud Purvis, Langley R Berman, Mervyn C Fibrin Fibrin - analysis The D-domain of fibrin (ogen) was separated from the parent molecule by plasmin digestion in the presence of calcium and isolated by means of DEAE-anion exchange chromatography followed by gel-filtration in buffer containing 4 M urea. Fluorescent-D-dimer (f-D-dimer) was isolated from a plasmic digest of fibrin clotted in the presence of 2.45 mM dansyl cadaverine, a fluorescent lysine analogue. Fluorescent-D-monomer was a by-product of f-D-dimer purification, the yield being determined by the concentration of dansyl cadaverine. At 2.45 mM f-D-monomer was always present in the digest. The f-D-monomer is probably formed directly and not as a result of degradation of f-D-dimer. The molecule elutes in the fibrinogen-derived-D- monomer position on gel-filtration. A protease was isolated and partially purified from venom of the puffadder (Bitis arietans). Puffadder venom protease is characterized by its ability to cleave D-dimer into symmetrical D-monomers, smaller than plasmin-derived D-monomers from fibrinogen. This characteristic was used to detect the puffadder venom protease activity with fluorescent-D-dimer being used as the substrate. Fractions obtained were assayed for D-dimer cleavage activity and the samples analyzed by means of SDS-PAGE on 4-20% gradient gels under reducing (βME) and non-reducing conditions. The fluorescent bands were located under U.V light and photographed prior to staining with Coomassie Blue. Several methods for the purification of the protease were investigated. 2018-02-01T13:29:57Z 2018-02-01T13:29:57Z 1987 Doctoral Thesis Doctoral PhD http://hdl.handle.net/11427/27202 eng application/pdf Division of Chemical Pathology Faculty of Health Sciences University of Cape Town |
| spellingShingle | Fibrin Fibrin - analysis Purves, Maud The D-domain of fibrin : structural and functional studies |
| thesis_degree_str | Doctoral |
| title | The D-domain of fibrin : structural and functional studies |
| title_full | The D-domain of fibrin : structural and functional studies |
| title_fullStr | The D-domain of fibrin : structural and functional studies |
| title_full_unstemmed | The D-domain of fibrin : structural and functional studies |
| title_short | The D-domain of fibrin : structural and functional studies |
| title_sort | d domain of fibrin structural and functional studies |
| topic | Fibrin Fibrin - analysis |
| url | http://hdl.handle.net/11427/27202 |
| work_keys_str_mv | AT purvesmaud theddomainoffibrinstructuralandfunctionalstudies AT purvesmaud ddomainoffibrinstructuralandfunctionalstudies |