RESUMEN
The present study describes the isolation and characterization of new pregnancy-associated glycoprotein molecules (PAG) from midpregnancy and late-pregnancy placentas in the water buffalo (Bubalus bubalis). After extraction, the homogenates are subjected to acid and ammonium sulfate precipitations followed by DEAE chromatography. Subsequently, the water buffalo PAG (wbPAG) from these solutions are enriched by Vicia villosa agarose (VVA) affinity chromatography. As determined by western blotting with anti-PAG sera, the apparent molecular masses of the immunoreactive bands from the VVA peaks range from 59.5 to 75.8kDa and from 57.8 to 73.3kDa in the midpregnancy and late-pregnancy placentas, respectively. Amino-terminal microsequencing of the immunoreactive proteins has allowed the identification of three distinct wbPAG sequences, which have been deposited in the SwissProt database: RGSXLTIHPLRNIRDFFYVG (acc. no. P85048), RGSXLTILPLRNIID (acc. no. P85049), and RGSXLTHLPLRNI (acc. no. P85050). Their comparison to previously identified proteins has shown that two of them are new because they have not been described before. Our results confirm the suitability of VVA chromatography for the enrichment of the multiple PAG molecules expressed in buffalo placenta.
Asunto(s)
Búfalos/fisiología , Placenta/química , Proteínas Gestacionales/aislamiento & purificación , Preñez/fisiología , Agar , Secuencia de Aminoácidos , Animales , Cromatografía de Afinidad , Cromatografía DEAE-Celulosa/métodos , Femenino , Desarrollo Fetal/fisiología , Fragmentos de Péptidos/química , Lectinas de Plantas , Embarazo , Proteínas Gestacionales/químicaRESUMEN
Placental tissue exhibits a typical glycosylation pattern, which differs from that observed in the pituitary gland. Depending to the species and pregnancy period, the placenta synthesizes diverse glycoproteins, some of which have significant hormonal activity, others being detected in maternal circulation. Thus, these molecules are of interest both from a fundamental and clinical point of view. Among the mammalian placental glycoproteins currently recognized, chorionic gonadotrophins from primates and Equidae, placental lactogen from bovines and the pregnancy-associated glycoproteins from ruminant species are particularly noteworthy. The diversity of saccharidic structures leads to multiple forms of placental glycoproteins exhibiting distinct structural and biological properties. For instance, concerning the chorionic gonadotrophins, the association of both alpha and beta subunits is essential for the binding of the hormone to specific receptors. Moreover, the N-linked oligossacharides are required for the activation of effectors systems. Bovine placental lactogen is a glycosylated hormone, exhibiting somatotropin- and prolactin-like activities. Several N-glycosylation sites confer to pregnancy-associated glycoproteins a long half-life (8-10 days) in maternal circulation. Assay of these molecules can be used for routine early pregnancy diagnosis and the follow-up of embryonic and fetal mortalities.