title: Multiple proteins from ejaculated and epididymal porcine spermatozoa bind glycan motifs found in the oviduct.
abstract: Previously, we demonstrated that porcine spermatozoa bind selectively to Lewis(x) (Le(X) ) and bi-antennary α2-6 sialylated N-acetyllactosamine (bi-SiaLN) glycan motifs found in the oviduct. Recognition of these motifs mediates sperm binding to oviduct epithelial cells to form a reservoir. Here, we characterize proteins that bind to Le(X) and bi-SiaLN in ejaculated and cauda epididymal spermatozoa. Fluoresceinated 3'-O-sulphated-Le(X) (suLe(X) ) and bi-SiaLN were used to locate the binding proteins on spermatozoa. Binding to N-acetyllactosamine (LN), a glycan that spermatozoa bind less, was assessed as a control. Over 60% of ejaculated and epididymal spermatozoa bound to suLe(X) and bi-SiaLN, but only 25% bound to LN. Specific binding to suLe(X) and bi-SiaLN required divalent cations. Fluoresceinated suLe(X) bound mostly to the plasma membrane at the apex of the head, whereas bi-SiaLN bound to a broader area of the head. Further characterization of receptors was accomplished by probing, with specific glycans, sperm proteins that had been separated by SDS-PAGE and transferred to blots. Glycan blot assays identified several suLe(X) -binding bands that migrated as 12-250 kDa. Migration of some of the bands that bound bi-SiaLN was similar to those that bound suLe(X) . A 12-kDa band protein was detected in seminal fluid and ejaculated spermatozoa, but absent in epididymal spermatozoa. In summary, we determined that: (i) multiple glycan-binding proteins recognize suLe(X) and bi-SiaLN motifs, (ii) specific binding required divalent cations and (iii) glycan-binding proteins were identified before and after sperm contact with accessory gland fluids. While the identity of these molecules is still unknown, this information predicts characteristics of the authentic oviduct suLe(X) and bi-SiaLN receptors on spermatozoa.
authors: Department of Animal Sciences, University of Illinois, Urbana, IL, USA.
authors: E
authors: E
authors: Silva
authors: G
authors: G
authors: Kadirvel
authors: R
authors: R
authors: Jiang
authors: N
authors: N
authors: Bovin
authors: D
authors: D
authors: Miller
mesh_terms: D000606:Amino Sugars / Q000378:metabolism; D000818:Animals; D004542:Ejaculation; D004823:Epididymitis; D005260:Female; D008297:Male; D010057:Oviducts / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D011485:Protein Binding; D029589:Seminal Plasma Proteins / Q000378:metabolism*; D013094:Spermatozoa / Q000378:metabolism*; D013552:Swine
chemical_list: D000606:Amino Sugars; D011134:Polysaccharides; D029589:Seminal Plasma Proteins; C032233:lactosamine; C000458:N-acetyllactosamine
keywords: Lewis X; carbohydrates; glycan-binding proteins; glycans; oviduct; sialic acid; spermatozoa