Structural, functional, and genetic characterization of Gastrophilus hemoglobin
Hemoglobin of Gastrophilus intestinalis (Insecta, Diptera), was purified and characterized. At least two isoforms have been identified by isoelectrofocusing, mass spectrometry, and genomic Southern blotting. Functional studies show a high oxygen affinity due to a low ligand dissociation rate (k(off)...
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ftunivgent:oai:archive.ugent.be:181007 2023-10-01T03:59:40+02:00 Structural, functional, and genetic characterization of Gastrophilus hemoglobin Dewilde, Sylvia Blaxter, Mark Van Hauwaert, Marie-Louise Van Houte, Koen Pesce, Alessandra Griffon, Nathalie Kiger, Laurent Marden, Michael C Vermeire, Sven Vanfleteren, Jacques Esmans, Eddy Moens, Luc 1998 application/pdf https://biblio.ugent.be/publication/181007 http://hdl.handle.net/1854/LU-181007 https://doi.org/10.1074/jbc.273.49.32467 https://biblio.ugent.be/publication/181007/file/3056306 eng eng https://biblio.ugent.be/publication/181007 http://hdl.handle.net/1854/LU-181007 http://dx.doi.org/10.1074/jbc.273.49.32467 https://biblio.ugent.be/publication/181007/file/3056306 No license (in copyright) info:eu-repo/semantics/restrictedAccess JOURNAL OF BIOLOGICAL CHEMISTRY ISSN: 0021-9258 Biology and Life Sciences INSECT CHIRONOMUS-THUMMI NONVERTEBRATE GLOBIN GENES AMINO-ACID SUBSTITUTION SPERM WHALE MYOGLOBIN LIGAND-BINDING ASCARIS HEMOGLOBIN INTRON POSITIONS CENTRAL EXON EVOLUTION SEQUENCES journalArticle info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion 1998 ftunivgent https://doi.org/10.1074/jbc.273.49.32467 2023-09-06T22:29:43Z Hemoglobin of Gastrophilus intestinalis (Insecta, Diptera), was purified and characterized. At least two isoforms have been identified by isoelectrofocusing, mass spectrometry, and genomic Southern blotting. Functional studies show a high oxygen affinity due to a low ligand dissociation rate (k(off) = 2.4 s(-1)) and a relatively high autoxidation rate (t(1/2) = 1.6/h), The globins were separated under denaturing conditions, and the sequence of Hb1 (M-r = 17,965 +/- 2) was determined at the protein and DNA level, The open reading frame codes for a polypeptide of 150 amino acids. Although the globin is distantly related to globins from other species, it has a low penalty score against globin templates. Freshly isolated hemoglobin was crystallized from polyethylene glycol. Crystals contain two hemoglobin molecules per asymmetric unit. Solution of the three-dimensional structure by molecular replacement could not be achieved, possibly due to the presence of three protein isoforms in the crystals. In order to determine its three-dimensional structure, G. intestinalis Hb1 was overexpressed in Escherichia coli, resulting in a fully functional molecule as confirmed by Ligand binding affinity. The globin gene contains two introns at positions D7.0 and G7.0. The D7.0 intron is unprecedented, suggesting that globin gene evolution is much more complex than originally thought. Article in Journal/Newspaper Sperm whale Ghent University Academic Bibliography Journal of Biological Chemistry 273 49 32467 32474 |
institution |
Open Polar |
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Ghent University Academic Bibliography |
op_collection_id |
ftunivgent |
language |
English |
topic |
Biology and Life Sciences INSECT CHIRONOMUS-THUMMI NONVERTEBRATE GLOBIN GENES AMINO-ACID SUBSTITUTION SPERM WHALE MYOGLOBIN LIGAND-BINDING ASCARIS HEMOGLOBIN INTRON POSITIONS CENTRAL EXON EVOLUTION SEQUENCES |
spellingShingle |
Biology and Life Sciences INSECT CHIRONOMUS-THUMMI NONVERTEBRATE GLOBIN GENES AMINO-ACID SUBSTITUTION SPERM WHALE MYOGLOBIN LIGAND-BINDING ASCARIS HEMOGLOBIN INTRON POSITIONS CENTRAL EXON EVOLUTION SEQUENCES Dewilde, Sylvia Blaxter, Mark Van Hauwaert, Marie-Louise Van Houte, Koen Pesce, Alessandra Griffon, Nathalie Kiger, Laurent Marden, Michael C Vermeire, Sven Vanfleteren, Jacques Esmans, Eddy Moens, Luc Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
topic_facet |
Biology and Life Sciences INSECT CHIRONOMUS-THUMMI NONVERTEBRATE GLOBIN GENES AMINO-ACID SUBSTITUTION SPERM WHALE MYOGLOBIN LIGAND-BINDING ASCARIS HEMOGLOBIN INTRON POSITIONS CENTRAL EXON EVOLUTION SEQUENCES |
description |
Hemoglobin of Gastrophilus intestinalis (Insecta, Diptera), was purified and characterized. At least two isoforms have been identified by isoelectrofocusing, mass spectrometry, and genomic Southern blotting. Functional studies show a high oxygen affinity due to a low ligand dissociation rate (k(off) = 2.4 s(-1)) and a relatively high autoxidation rate (t(1/2) = 1.6/h), The globins were separated under denaturing conditions, and the sequence of Hb1 (M-r = 17,965 +/- 2) was determined at the protein and DNA level, The open reading frame codes for a polypeptide of 150 amino acids. Although the globin is distantly related to globins from other species, it has a low penalty score against globin templates. Freshly isolated hemoglobin was crystallized from polyethylene glycol. Crystals contain two hemoglobin molecules per asymmetric unit. Solution of the three-dimensional structure by molecular replacement could not be achieved, possibly due to the presence of three protein isoforms in the crystals. In order to determine its three-dimensional structure, G. intestinalis Hb1 was overexpressed in Escherichia coli, resulting in a fully functional molecule as confirmed by Ligand binding affinity. The globin gene contains two introns at positions D7.0 and G7.0. The D7.0 intron is unprecedented, suggesting that globin gene evolution is much more complex than originally thought. |
format |
Article in Journal/Newspaper |
author |
Dewilde, Sylvia Blaxter, Mark Van Hauwaert, Marie-Louise Van Houte, Koen Pesce, Alessandra Griffon, Nathalie Kiger, Laurent Marden, Michael C Vermeire, Sven Vanfleteren, Jacques Esmans, Eddy Moens, Luc |
author_facet |
Dewilde, Sylvia Blaxter, Mark Van Hauwaert, Marie-Louise Van Houte, Koen Pesce, Alessandra Griffon, Nathalie Kiger, Laurent Marden, Michael C Vermeire, Sven Vanfleteren, Jacques Esmans, Eddy Moens, Luc |
author_sort |
Dewilde, Sylvia |
title |
Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
title_short |
Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
title_full |
Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
title_fullStr |
Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
title_full_unstemmed |
Structural, functional, and genetic characterization of Gastrophilus hemoglobin |
title_sort |
structural, functional, and genetic characterization of gastrophilus hemoglobin |
publishDate |
1998 |
url |
https://biblio.ugent.be/publication/181007 http://hdl.handle.net/1854/LU-181007 https://doi.org/10.1074/jbc.273.49.32467 https://biblio.ugent.be/publication/181007/file/3056306 |
genre |
Sperm whale |
genre_facet |
Sperm whale |
op_source |
JOURNAL OF BIOLOGICAL CHEMISTRY ISSN: 0021-9258 |
op_relation |
https://biblio.ugent.be/publication/181007 http://hdl.handle.net/1854/LU-181007 http://dx.doi.org/10.1074/jbc.273.49.32467 https://biblio.ugent.be/publication/181007/file/3056306 |
op_rights |
No license (in copyright) info:eu-repo/semantics/restrictedAccess |
op_doi |
https://doi.org/10.1074/jbc.273.49.32467 |
container_title |
Journal of Biological Chemistry |
container_volume |
273 |
container_issue |
49 |
container_start_page |
32467 |
op_container_end_page |
32474 |
_version_ |
1778533964616564736 |