Primary structure of kappa-casein isolated from mares' milk.

In this work the purification and the complete primary structure of kappa-casein from equine milk are reported for the first time. Mares' milk casein was separated by RP-HPLC into four fractions. Complete primary sequence was obtained by sequence analysis of the protein in the fastest eluting peak isolated by chromatography. This sequence was 95% identical to that reported for the C-terminal portion of the zebras' kappa-casein and showed high similarity with kappa-caseins from sources other than Equidae, confirming that this protein was indeed kappa-casein in equine milk. The presence of post-translational modifications in equine kappa-casein was investigated by mass spectroscopy, after enzymic dephosphorylation. Two main components were found, the smaller component being more abundant. Equine kappa-casein was recognized by a lectin specific for one of the glucosidic bonds in the saccharide moiety of bovine kappa-casein. Sequence comparison with prevision studies showed that the distribution of charged and hydrophobic regions in equine kappa-casein was similar, but not identical, to that found in the bovine protein; these regions are associated with the role of kappa-casein in the formation and stabilization of the micellar structure of casein in milk.

Purification and primary structure of a new bovine spermadhesin.

Z13 is a new seminal plasma protein made up of two disulfide-linked 13-kDa subunits that was identified in our laboratory by 2D PAGE. In this report we present the purification of Z13 from bovine seminal plasma. In solution, the protein is a nonglycosylated dimer that presents one interchain disulfide bond and does not show heparin-binding properties. The complete primary structure and the localization of the S-S bridges are reported. The results suggest that Z13 is a new protein of the spermadhesin family whose members are thought to play a prominent role in different aspects of fertilization.

D-aspartate oxidase is present in ovaries, eggs and embryos but not in testis of Xenopus laevis.

D-aspartate oxidase (DASPO) is an FAD dependent flavoprotein which catalyzes the oxidative deamination of D-aspartate using oxygen as electron acceptor. D-aspartate and DASPO are supposed to be involved in the regulation of the central nervous system and in the animal development. This manuscript describes for the first time the presence of DASPO in Xenopus laevis fertilized eggs and embryos and suggests a different tissue distribution of this enzyme in adult male and female animals. In particular, by means of 2D-electrophoresis and affinity purified specific anti-DASPO antibodies, the enzyme was localized in fertilized eggs of X. laevis and in ovaries of adult animals but it was shown to be absent in the testis suggesting a gender specific expression. The protein from Xenopus ovaries has been purified by means of immunoprecipitation and it has M(r) of 30 kDa and pI of 8.1.