Identification and characterization of non-phosphorylcholine-binding and phosphorylcholine-binding proteins of canine seminal plasma

Seminal plasma (SP) proteins participate in the process of fertilization by binding to the sperm membrane, particularly to the phosphorylcholine-containing lipids. This study aimed to identify and characterize non-phosphorylcholine-binding and phosphorylcholine-binding proteins (nPch- BPs and PchBPs, respectively) of canine SP. The nPchBPs and PchBPs were isolated from canine SP by affinity chromatography. Electrophoretic studies revealed that the nPchBPs and PchBPs occurred in their native state as high-molecular-weight aggregates. Immunofluorescent staining showed preferential binding of nPchBPs to the sperm acrosome membrane, whereas PchBPs coating was uniformly distributed on the sperm post-acrosomal membrane, mid-piece and tail regions. Analysis with mass spectrometry confirmed that canine prostate specific esterase (CPSE) is a component of the nPchBPs and PchBPs, which is implicated in key mechanisms of protein-coating on the sperm plasma membrane surface. In addition, proteins of known binding properties such as prostaglandin-H2 D-isomerase and lipocalin-like 1 protein, identified in canine SP, might have a specific role in the fertilization-associated processes.