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Interaction of milk proteins and Binder of Sperm (BSP) proteins from boar, stallion and ram semen.

Plante G, Lusignan MF, Lafleur M, Manjunath P - Reprod. Biol. Endocrinol. (2015)

Bottom Line: These changes can ultimately be detrimental to sperm storage.They also had affinity for another milk protein fraction (F2) composed mostly of casein micelles.However, stallion BSP showed higher affinity for the fraction (F1).

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Université de Montréal, C.P. 6128, Succ. Centre Ville, Montréal, Québec, Canada, H3C 3J7. genevieve.plante@umontreal.ca.

ABSTRACT

Background: Mammalian semen contains a family of closely related proteins known as Binder of SPerm (BSP proteins) that are added to sperm at ejaculation. BSP proteins extract lipids from the sperm membrane thereby extensively modifying its composition. These changes can ultimately be detrimental to sperm storage. We have demonstrated that bovine BSP proteins interact with major milk proteins and proposed that this interaction could be the basis of sperm protection by milk extenders. In the present study, we investigated if homologous BSP proteins present in boar, stallion and ram seminal plasma display a similar affinity for the milk proteins in order to assess whether the mechanism of sperm protection by milk for these species could be general.

Methods: Skim milk was incubated with seminal plasma proteins (boar, stallion and ram), chromatographed on a Sepharose CL-4B column and protein fractions were analyzed by immunoblotting.

Results: Boar, stallion and ram BSP proteins displayed affinity for a milk protein fraction (F1) mainly composed of α-lactalbumin, β-lactoglobulin, and κ-casein. They also had affinity for another milk protein fraction (F2) composed mostly of casein micelles. However, stallion BSP showed higher affinity for the fraction (F1).

Conclusions: These results further extend our view that the association of BSP proteins with milk proteins could be a general feature of the mechanism of mammalian sperm protection by milk to prevent detrimental effect of prolonged exposure of sperm to seminal plasma.

No MeSH data available.


Related in: MedlinePlus

Isolation and identification of ram BSP proteins. a Chromatogram of ram SP proteins (150 mg) on gelatin-agarose column. The gelatin-adsorbed (GA) proteins (tubes 55–61) were pooled, desalted and freeze-dried. b SDS-PAGE of ram SP proteins (20 μg) and of GA fraction (15 μg). c Immunoblot analysis of ram BSP proteins using the polyclonal antibodies directed against ram BSP proteins. Ram SP proteins (2 μg) and GA proteins (200 ng). d Specificity of the polyclonal antibodies directed against ram BSP proteins. SP, ram seminal plasma proteins (300 ng) and SM, skimmed milk proteins (10 μg)
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Fig2: Isolation and identification of ram BSP proteins. a Chromatogram of ram SP proteins (150 mg) on gelatin-agarose column. The gelatin-adsorbed (GA) proteins (tubes 55–61) were pooled, desalted and freeze-dried. b SDS-PAGE of ram SP proteins (20 μg) and of GA fraction (15 μg). c Immunoblot analysis of ram BSP proteins using the polyclonal antibodies directed against ram BSP proteins. Ram SP proteins (2 μg) and GA proteins (200 ng). d Specificity of the polyclonal antibodies directed against ram BSP proteins. SP, ram seminal plasma proteins (300 ng) and SM, skimmed milk proteins (10 μg)

Mentions: The same approach was used to produce polyclonal antibodies against ram BSP proteins. Purification of ram BSP proteins has already been described [20]. As previously reported, a large fraction of ram SP proteins bound specifically to the gelatin-agarose column (Fig. 2a). Gelatin-agarose bound proteins were separated by SDS-PAGE and bands of 15–16 kDa and 22–24 kDa, previously shown to be BSP proteins, were observed (Fig. 2b). Antibodies raised against the gelatin-binding proteins specifically recognized the 15–16 kDa and 22–24 kDa BSP proteins (Fig. 2c). They did not recognize any of the milk proteins (Fig. 2d).Fig. 2


Interaction of milk proteins and Binder of Sperm (BSP) proteins from boar, stallion and ram semen.

Plante G, Lusignan MF, Lafleur M, Manjunath P - Reprod. Biol. Endocrinol. (2015)

Isolation and identification of ram BSP proteins. a Chromatogram of ram SP proteins (150 mg) on gelatin-agarose column. The gelatin-adsorbed (GA) proteins (tubes 55–61) were pooled, desalted and freeze-dried. b SDS-PAGE of ram SP proteins (20 μg) and of GA fraction (15 μg). c Immunoblot analysis of ram BSP proteins using the polyclonal antibodies directed against ram BSP proteins. Ram SP proteins (2 μg) and GA proteins (200 ng). d Specificity of the polyclonal antibodies directed against ram BSP proteins. SP, ram seminal plasma proteins (300 ng) and SM, skimmed milk proteins (10 μg)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4536704&req=5

Fig2: Isolation and identification of ram BSP proteins. a Chromatogram of ram SP proteins (150 mg) on gelatin-agarose column. The gelatin-adsorbed (GA) proteins (tubes 55–61) were pooled, desalted and freeze-dried. b SDS-PAGE of ram SP proteins (20 μg) and of GA fraction (15 μg). c Immunoblot analysis of ram BSP proteins using the polyclonal antibodies directed against ram BSP proteins. Ram SP proteins (2 μg) and GA proteins (200 ng). d Specificity of the polyclonal antibodies directed against ram BSP proteins. SP, ram seminal plasma proteins (300 ng) and SM, skimmed milk proteins (10 μg)
Mentions: The same approach was used to produce polyclonal antibodies against ram BSP proteins. Purification of ram BSP proteins has already been described [20]. As previously reported, a large fraction of ram SP proteins bound specifically to the gelatin-agarose column (Fig. 2a). Gelatin-agarose bound proteins were separated by SDS-PAGE and bands of 15–16 kDa and 22–24 kDa, previously shown to be BSP proteins, were observed (Fig. 2b). Antibodies raised against the gelatin-binding proteins specifically recognized the 15–16 kDa and 22–24 kDa BSP proteins (Fig. 2c). They did not recognize any of the milk proteins (Fig. 2d).Fig. 2

Bottom Line: These changes can ultimately be detrimental to sperm storage.They also had affinity for another milk protein fraction (F2) composed mostly of casein micelles.However, stallion BSP showed higher affinity for the fraction (F1).

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Université de Montréal, C.P. 6128, Succ. Centre Ville, Montréal, Québec, Canada, H3C 3J7. genevieve.plante@umontreal.ca.

ABSTRACT

Background: Mammalian semen contains a family of closely related proteins known as Binder of SPerm (BSP proteins) that are added to sperm at ejaculation. BSP proteins extract lipids from the sperm membrane thereby extensively modifying its composition. These changes can ultimately be detrimental to sperm storage. We have demonstrated that bovine BSP proteins interact with major milk proteins and proposed that this interaction could be the basis of sperm protection by milk extenders. In the present study, we investigated if homologous BSP proteins present in boar, stallion and ram seminal plasma display a similar affinity for the milk proteins in order to assess whether the mechanism of sperm protection by milk for these species could be general.

Methods: Skim milk was incubated with seminal plasma proteins (boar, stallion and ram), chromatographed on a Sepharose CL-4B column and protein fractions were analyzed by immunoblotting.

Results: Boar, stallion and ram BSP proteins displayed affinity for a milk protein fraction (F1) mainly composed of α-lactalbumin, β-lactoglobulin, and κ-casein. They also had affinity for another milk protein fraction (F2) composed mostly of casein micelles. However, stallion BSP showed higher affinity for the fraction (F1).

Conclusions: These results further extend our view that the association of BSP proteins with milk proteins could be a general feature of the mechanism of mammalian sperm protection by milk to prevent detrimental effect of prolonged exposure of sperm to seminal plasma.

No MeSH data available.


Related in: MedlinePlus