Identificación de residuos de carbohidratos de proteínas asociadas a balsas lipídicas (rafts) de espermatozoides de cerdo
Mammalian sperm are motile after ejaculation, but cannot fertilize the egg. This ability is acquired during passage through the female reproductive tract and it is called sperm capacitation. In this process the activation of both, a bicarbonate (HCO₃- ) dependent adenylate cyclase and a protein kina...
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| Tipo de recurso: | tesis de maestría |
| Estado: | Versión publicada |
| Fecha de publicación: | 2011 |
| País: | México |
| Institución: | Universidad Autónoma Metropolitana |
| Repositorio: | Repositorio Institucional de la UAM Iztapalapa |
| Idioma: | español |
| OAI Identifier: | oai:bindani.izt.uam.mx:8w32r580d |
| Acceso en línea: | https://doi.org/10.24275/uami.8w32r580d |
| Access Level: | acceso abierto |
| Palabra clave: | info:eu-repo/classification/LEM/Espermatozoides info:eu-repo/classification/LEM/Spermatozoa info:eu-repo/classification/LEM/Cerdo -- Reproducción info:eu-repo/classification/LEM/Biología experimental info:eu-repo/classification/LEM/Swine -- Reproduction info:eu-repo/classification/LEM/Biology, Experimental info:eu-repo/classification/cti/3 |
| Sumario: | Mammalian sperm are motile after ejaculation, but cannot fertilize the egg. This ability is acquired during passage through the female reproductive tract and it is called sperm capacitation. In this process the activation of both, a bicarbonate (HCO₃- ) dependent adenylate cyclase and a protein kinase A (PKA) allow tyrosine phosphorylation of a group of proteins, and this is associated with changes in sperm motility and acrosome reaction. The plasma membrane is modified during the capacitation, especially lipid rafts, small platforms within the plasma membrane, composed of specific proteins, glycosphingolipids, gangliosides and cholesterol in the exoplasmic face, connected to phospholipids and cholesterol on the cytoplasmic face. The signaling proteins of the lipid rafts are specific of the cell type and physiological state, and include non-receptor and receptor proteins with tyrosine kinase activity, G proteins, inositol phospholipids, glycosylphosphatidylinositol (GPI)-anchored proteins, nitric oxide synthase and others. The aim of this work was to identify carbohydrate residues of proteins associated with lipid rafts of fresh and capacitated boar spermatozoa. Seven semen samples of healthy boars were analyzed. Capacitation was induced in vitro by incubating in TALP-Hepes medium during 4 hours at 39 ˚C. Lipid rafts were obtained by ultracentrifugation in discontinuous sucrose gradient, specifically in the 5-35% interphase, in both conditions. Electrophoresis was performed with 12.5 % polyacrylamide gels, with 20 g of protein per lane. The fraction corresponding to lipid rafts of fresh sperm showed 16 bands with Mr of 11 to 134 kDa and in capacitated sperm, from 11 to 141 kDa. Carbohydrate residues identification was performed by Western blot, using DSA and SNA lectins, specific for N-acetylglucosamine and sialic acid, respectively. DSA lectin joined to 8 bands of proteins in rafts of fresh sperm and 11 of capacitated sperm. SNA joined 4 bands in fresh sperm and 7 in the capacitated. Our results show that the proteins associated with lipid rafts of capacitated spermatozoa are glycosylated with residues of Nacetylglucosamine and sialic acid, and the number of glycosylated proteins associated with lipid rafts increases after capacitation. |
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