Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts

Autores
Zahn, Astrid; Furlong, Laura Ines; Biancotti, Juan Carlos; Ghiringhelli, Pablo Daniel; Marin Briggiler, Clara Isabel; Vazquez, Monica Hebe
Año de publicación
2002
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Acrosin is an acrosomal protease synthesized as a proenzyme and activated into beta-acrosin during the acrosome reaction. In the present study, a set of sensitive assays was developed to identify the proacrosin/acrosin system and to evaluate its activation pattern in human sperm extracts. Immunocytochemical analysis with monoclonal antibody (Mab) AcrC5F10 showed specific staining on the acrosome of permeabilized ejaculated and capacitated spermatozoa. Acrosome reaction was associated with a decrease in staining. AcrC5F10 specifically recognized a 55-kDa band (proacrosin) in Western immunoblots. Activation studies showed enzymatically active intermediates of 39 and 35 kDa after zymography. Immunoreactive bands of 52, 43, 34, 21-26 and 16 kDa were identified in the activation patterns developed with AcrC5F10. Activation was completely inhibited in the presence of 9 mM CaCl(2) or 100 mM benzamidine. A multiple sequence alignment revealed partial conservation of putative cleavage sites in the proacrosin sequence. The tests described allow the detection of human proacrosin in spermatozoa and sperm protein extracts, as well as the evaluation of the proenzyme activation pattern. They can be used to study the effect of inhibitors upon proenzyme activation. In addition, alterations in proacrosin activation in semen samples with abnormal acrosin enzymatic activity can be analyzed using these assays.
Fil: Zahn, Astrid. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Furlong, Laura Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Biancotti, Juan Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Ghiringhelli, Pablo Daniel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; Argentina
Fil: Marin Briggiler, Clara Isabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Vazquez, Monica Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Materia
Human Spermatozoa
Proteases
Acrosin
Activation
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/31329
Acceder
id CONICETDig_d667de7802802ab2930e36cfe78d85cc
oai_identifier_str oai:ri.conicet.gov.ar:11336/31329
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extractsZahn, AstridFurlong, Laura InesBiancotti, Juan CarlosGhiringhelli, Pablo DanielMarin Briggiler, Clara IsabelVazquez, Monica HebeHuman SpermatozoaProteasesAcrosinActivationhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Acrosin is an acrosomal protease synthesized as a proenzyme and activated into beta-acrosin during the acrosome reaction. In the present study, a set of sensitive assays was developed to identify the proacrosin/acrosin system and to evaluate its activation pattern in human sperm extracts. Immunocytochemical analysis with monoclonal antibody (Mab) AcrC5F10 showed specific staining on the acrosome of permeabilized ejaculated and capacitated spermatozoa. Acrosome reaction was associated with a decrease in staining. AcrC5F10 specifically recognized a 55-kDa band (proacrosin) in Western immunoblots. Activation studies showed enzymatically active intermediates of 39 and 35 kDa after zymography. Immunoreactive bands of 52, 43, 34, 21-26 and 16 kDa were identified in the activation patterns developed with AcrC5F10. Activation was completely inhibited in the presence of 9 mM CaCl(2) or 100 mM benzamidine. A multiple sequence alignment revealed partial conservation of putative cleavage sites in the proacrosin sequence. The tests described allow the detection of human proacrosin in spermatozoa and sperm protein extracts, as well as the evaluation of the proenzyme activation pattern. They can be used to study the effect of inhibitors upon proenzyme activation. In addition, alterations in proacrosin activation in semen samples with abnormal acrosin enzymatic activity can be analyzed using these assays.Fil: Zahn, Astrid. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Furlong, Laura Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Biancotti, Juan Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Ghiringhelli, Pablo Daniel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; ArgentinaFil: Marin Briggiler, Clara Isabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaFil: Vazquez, Monica Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; ArgentinaElsevier Ireland2002-03-02info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/31329Vazquez, Monica Hebe; Marin Briggiler, Clara Isabel; Ghiringhelli, Pablo Daniel; Biancotti, Juan Carlos; Furlong, Laura Ines; Zahn, Astrid; et al.; Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts; Elsevier Ireland; Journal Of Reproductive Immunology; 54; 1-2; 2-3-2002; 43-630165-03781872-7603CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0165037801000808?via%3Dihubinfo:eu-repo/semantics/altIdentifier/doi/10.1016/S0165-0378(01)00080-8info:eu-repo/semantics/altIdentifier/pmid/https://www.ncbi.nlm.nih.gov/pubmed/11839395info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-nd/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:44:10Zoai:ri.conicet.gov.ar:11336/31329instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 10:44:10.733CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
title Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
spellingShingle Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
Zahn, Astrid
Human Spermatozoa
Proteases
Acrosin
Activation
title_short Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
title_full Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
title_fullStr Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
title_full_unstemmed Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
title_sort Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts
dc.creator.none.fl_str_mv Zahn, Astrid
Furlong, Laura Ines
Biancotti, Juan Carlos
Ghiringhelli, Pablo Daniel
Marin Briggiler, Clara Isabel
Vazquez, Monica Hebe
author Zahn, Astrid
author_facet Zahn, Astrid
Furlong, Laura Ines
Biancotti, Juan Carlos
Ghiringhelli, Pablo Daniel
Marin Briggiler, Clara Isabel
Vazquez, Monica Hebe
author_role author
author2 Furlong, Laura Ines
Biancotti, Juan Carlos
Ghiringhelli, Pablo Daniel
Marin Briggiler, Clara Isabel
Vazquez, Monica Hebe
author2_role author
author
author
author
author
dc.subject.none.fl_str_mv Human Spermatozoa
Proteases
Acrosin
Activation
topic Human Spermatozoa
Proteases
Acrosin
Activation
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Acrosin is an acrosomal protease synthesized as a proenzyme and activated into beta-acrosin during the acrosome reaction. In the present study, a set of sensitive assays was developed to identify the proacrosin/acrosin system and to evaluate its activation pattern in human sperm extracts. Immunocytochemical analysis with monoclonal antibody (Mab) AcrC5F10 showed specific staining on the acrosome of permeabilized ejaculated and capacitated spermatozoa. Acrosome reaction was associated with a decrease in staining. AcrC5F10 specifically recognized a 55-kDa band (proacrosin) in Western immunoblots. Activation studies showed enzymatically active intermediates of 39 and 35 kDa after zymography. Immunoreactive bands of 52, 43, 34, 21-26 and 16 kDa were identified in the activation patterns developed with AcrC5F10. Activation was completely inhibited in the presence of 9 mM CaCl(2) or 100 mM benzamidine. A multiple sequence alignment revealed partial conservation of putative cleavage sites in the proacrosin sequence. The tests described allow the detection of human proacrosin in spermatozoa and sperm protein extracts, as well as the evaluation of the proenzyme activation pattern. They can be used to study the effect of inhibitors upon proenzyme activation. In addition, alterations in proacrosin activation in semen samples with abnormal acrosin enzymatic activity can be analyzed using these assays.
Fil: Zahn, Astrid. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Furlong, Laura Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Biancotti, Juan Carlos. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Ghiringhelli, Pablo Daniel. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; Argentina
Fil: Marin Briggiler, Clara Isabel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
Fil: Vazquez, Monica Hebe. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental. Fundación de Instituto de Biología y Medicina Experimental. Instituto de Biología y Medicina Experimental; Argentina
description Acrosin is an acrosomal protease synthesized as a proenzyme and activated into beta-acrosin during the acrosome reaction. In the present study, a set of sensitive assays was developed to identify the proacrosin/acrosin system and to evaluate its activation pattern in human sperm extracts. Immunocytochemical analysis with monoclonal antibody (Mab) AcrC5F10 showed specific staining on the acrosome of permeabilized ejaculated and capacitated spermatozoa. Acrosome reaction was associated with a decrease in staining. AcrC5F10 specifically recognized a 55-kDa band (proacrosin) in Western immunoblots. Activation studies showed enzymatically active intermediates of 39 and 35 kDa after zymography. Immunoreactive bands of 52, 43, 34, 21-26 and 16 kDa were identified in the activation patterns developed with AcrC5F10. Activation was completely inhibited in the presence of 9 mM CaCl(2) or 100 mM benzamidine. A multiple sequence alignment revealed partial conservation of putative cleavage sites in the proacrosin sequence. The tests described allow the detection of human proacrosin in spermatozoa and sperm protein extracts, as well as the evaluation of the proenzyme activation pattern. They can be used to study the effect of inhibitors upon proenzyme activation. In addition, alterations in proacrosin activation in semen samples with abnormal acrosin enzymatic activity can be analyzed using these assays.
publishDate 2002
dc.date.none.fl_str_mv 2002-03-02
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/31329
Vazquez, Monica Hebe; Marin Briggiler, Clara Isabel; Ghiringhelli, Pablo Daniel; Biancotti, Juan Carlos; Furlong, Laura Ines; Zahn, Astrid; et al.; Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts; Elsevier Ireland; Journal Of Reproductive Immunology; 54; 1-2; 2-3-2002; 43-63
0165-0378
1872-7603
CONICET Digital
CONICET
url http://hdl.handle.net/11336/31329
identifier_str_mv Vazquez, Monica Hebe; Marin Briggiler, Clara Isabel; Ghiringhelli, Pablo Daniel; Biancotti, Juan Carlos; Furlong, Laura Ines; Zahn, Astrid; et al.; Evaluation of the proacrosin / acrosin system and its mechanism of activation in human sperm extracts; Elsevier Ireland; Journal Of Reproductive Immunology; 54; 1-2; 2-3-2002; 43-63
0165-0378
1872-7603
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/http://www.sciencedirect.com/science/article/pii/S0165037801000808?via%3Dihub
info:eu-repo/semantics/altIdentifier/doi/10.1016/S0165-0378(01)00080-8
info:eu-repo/semantics/altIdentifier/pmid/https://www.ncbi.nlm.nih.gov/pubmed/11839395
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-nd/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Elsevier Ireland
publisher.none.fl_str_mv Elsevier Ireland
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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score 13.070432

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