Preparation of human tissue protein extracts enriched with the sphingomyelin synthase 1

Sphingomyelin synthase 1 (SMS1) catalyzes sphingomyelin biosynthesis in eukaryotic cells. We previously studied the structure of the human SGMS1 gene, which encodes the enzyme and its numerous transcripts. The tissue-specific expression of the transcripts was also described. Analysis of the SMS1 protein expression in human tissues using immunoblotting of tissue extracts prepared in the RIPA (Radio Immuno- Precipitation Assay) buffer revealed a weak signal in the renal cortex, testis, lung, and no signal in the placenta or lymphatic node. In this work, a new method of the preparation of the tissue protein extracts enriched with SMS1 was suggested. The method based on the consecutive extraction with a buffer containing 0.05 and 1 mg/mL of the Quillaja saponaria saponin allowed SMS1 to be detected in all tissues tested. The SMS1 con- tent in the saponin extract of kidney cortex is about 12-fold higher than in the RIPA extraction procedure.