Characterization, expression and antibacterial properties of apolipoproteins A from carp (Cyprinus carpio L.) seminal plasma

Abstract

Apolipoproteins A are multifunctional proteins that, in addition to contributing to lipid metabolism and transport, are associated with the innate immune system in fish. Using a three step isolation procedure consisting of affinity chromatography on Blue-Sepharose, delipidation and reverse phase HPLC we isolated apolipoproteins from carp seminal plasma and identified them as ApoA-I and Apo-14 kDa. Moreover, we provided the full-length cDNA sequence of ApoA-I encoding 257 amino acids including a 18 amino acid signal peptide and a 4 amino acid propeptide. Apolipoproteins corresponded to the most abundant proteins in carp seminal plasma. Both ApoA-I and Apo-14 kDa were represented by several proteoforms that differ both in molecular mass and isoelectric point. The proteoforms of ApoA-I characteristic for seminal plasma were distinguished from those of blood. Carp seminal plasma ApoA-I and Apo-14 kDa showed a high immunologic similarity to their counterparts in carp blood and seminal plasma of other Cyprinid species. The mRNA expression analysis and immunohistochemical study suggest synthesis and secretion of ApoA-I and Apo-14 kDa in the fish reproductive tract and suggest a role in spermatogenesis and the stabilization of sperm membrane. Moreover, ApoA-I displayed bactericidal activity against Escherichia coli and bacteriostatic activity against Aeromonas hydrophila which suggests that ApoA-I is associated with innate immune system of the fish reproductive tract.