Solubilization and affinity chromatography of a sialidase from human liver.
A sialidase, acting on gangliosides and mucus glycoproteins (pH optimum 4.0-4.5) was solubilized by 1% Triton X-100 and short ultrasonication from a crude mitochondrial-lysosomal fraction isolated from human liver. The enzyme was enriched over 1000-fold with the aid of affinity chromatography on equine submandibular gland mucin bound to Sepharose and 20mM Tris/HCl buffer, pH 7.5, as eluent. The sialidase exhibited a molecular mass of about 200 000 Da on Sephadex G-200. On analytical gel electrophoresis, an enzymically active protein band of about 70 000 Da was observed. A sialidase acting on sialyllactose remained in the membrane fraction and could not be solubilized.
D002846:Chromatography, Affinity / Q000379:methods; D005732:Gangliosides; D006023:Glycoproteins; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D009439:Neuraminidase / Q000302:isolation & purification* / Q000378:metabolism; D012995:Solubility; D013379:Substrate Specificity
D005732:Gangliosides; D006023:Glycoproteins; D009439:Neuraminidase