Research

The solubilisation of boar sperm membranes by different detergents - a microscopic, MALDI-TOF MS, ³¹P NMR and PAGE study on membrane lysis, extraction efficiency, lipid and protein composition

Ulrike Jakop^1* , Beate Fuchs^2* , Rosmarie Süß² , Gudrun Wibbelt¹ , Beate Braun¹ , Karin Müller^1* and Jürgen Schiller²

¹  Leibniz Institute for Zoo and Wildlife Research, Alfred-Kowalke-Str. 17, D-10315 Berlin, Germany

²  University of Leipzig, Faculty of Medicine, Institute of Medical Physics and Biophysics, Härtelstr. 16-18, D-04107 Leipzig, Germany

author email corresponding author email* Contributed equally

Lipids in Health and Disease 2009, 8:49doi:10.1186/1476-511X-8-49

Published:	11 November 2009

Abstract

Background

Detergents are often used to isolate proteins, lipids as well as "detergent-resistant membrane domains" (DRMs) from cells. Different detergents affect different membrane structures according to their physico-chemical properties. However, the effects of different detergents on membrane lysis of boar spermatozoa and the lipid composition of DRMs prepared from the affected sperm membranes have not been investigated so far.

Results

Spermatozoa were treated with the selected detergents Pluronic F-127, sodium cholate, CHAPS, Tween 20, Triton X-100 and Brij 96V. Different patterns of membrane disintegration were observed by light and electron microscopy. In accordance with microscopic data, different amounts of lipids and proteins were released from the cells by the different detergents. The biochemical methods to assay the phosphorus and cholesterol contents as well as ³¹P NMR to determine the phospholipids were not influenced by the presence of detergents since comparable amounts of lipids were detected in the organic extracts from whole cell suspensions after exposure to each detergent. However, matrix-assisted laser desorption and ionization time-of-flight mass spectrometry applied to identify phospholipids was essentially disturbed by the presence of detergents which exerted particular suppression effects on signal intensities. After separation of the membrane fractions released by detergents on a sucrose gradient only Triton X-100 and sodium cholate produced sharp turbid DRM bands. Only membrane solubilisation by Triton X-100 leads to an enrichment of cholesterol, sphingomyelin, phosphatidylinositol and phosphatidylethanolamine in a visible DRM band accompanied by a selective accumulation of proteins.

Conclusion

The boar sperm membranes are solubilised to a different extent by the used detergents. Particularly, the very unique DRMs isolated after Triton X-100 exposure are interesting candidates for further studies regarding the architecture of sperm.