Carbohydrate composition of purified human liver ?-L-Fucosidase

Abstract

Human liver alpha-L-fucosidase was purified to apparent homogeneity and analyzed for carbohydrate content primarily by gas-liquid chromatography (glc). The enzyme is about 7% carbohydrate by weight and contains the following sugars (residues per 50, 000 molecular weight subunit): mannose (8.3), glucosamine (4.3) (presumably N-acetylated), sialic acid (1.6) and glucose (1.6). Galactose (0.8) and L-fucose (1.8) were also found but their presence may be due to artifacts of the purification procedure.

Structure and Substrate Specificity of a Eukaryotic Fucosidase from Fusarium graminearum

“…We found the total carbohydrate content of FgFCO1 expressed from P. pastoris to be ϳ6% based on the major peak identified by mass spectrometry. Human fucosidases FUCA1 and FUCA2 are also known to be N-glycosylated (66,67). The liver fucosidase was reported to contain Man, GlcNAc, sialic acid, and Glc contents with a total of ϳ7% carbohydrates by weight (67).…”

Carbohydrate composition of human placental N-acetylhexosaminidase A and B

“…Samples (2-5,ul) of the derivatives containing the equivalent of 12-40,g of original enzyme protein were injected into the gas chromatograph and analysed as previously described, except that the oven temperature increase was programmed at 2°C/min (Alhadeff& Freeze, 1977). To determine the reliability ofthe analyses ofvery small amounts ofcarbohydrate, samples containing 20-200,ug of ovalbumin (1-10ug of total carbohydrate) and 0.5-5.0,ug of arabitol were similarly analysed by following identical sample preparations and derivative-preparation procedures.…”

Purification and characterization of altered cystic fibrosis liver α‐L‐fucosidase

“…Carbohydrate analyses. Carbohydrates were analyzed as previously described for purified normal liver a-L-fucosidase (Alhadeff & Freeze 1977) by gas-liquid chromato-graphy (glc) using trimethylsilyl (TMS) derivatives of the methyl glycosides, essentially as described by Etchison & Holland (1974). Samples containing 140-300 pg of enzyme protein (Lowry) were lyophilized in screw-cap vials containing 5 pg of arabitol (internal standard) and the samples were methanolyzed with 0.5 ml of 1.5 N anhydrous methanolic-HC1, prepared according to Clamp et al (1973).…”

“…In a previous study, altered a-Lfucosidase isoenzyme patterns were found in cystic fibrosis livers (Alhadeff et al 1976). Since there is considerable evidence that liver a-L-fucosidase is a sialoglycoprotein (Alhadeff & Freeze 1977) and that the isoenzymes are related by sialic acid residues (Alhadeff et al 1974, 1975b, Thorpe & Robinson 1975, we suggested that the altered fucosidase profiles found in cystic fibrosis might result from aberrant glycosylation (Alhadeff et al 1976). In a recent study we have investigated the activity and properties of glycoprotein-sialyltransferase in cystic fibrosis livers (Alhadeff & Cimino 1978).…”

“…In an attempt to understand better the altered a-L-fucosidase profiles found in cystic fibrosis livers and to provide more direct evidence for defective glycosylation, we purified and characterized a-l-fucosidases from two cystic fibrosis livers and compared their properties to those of the purified enzyme from normal liver (Alhadeff et al 1975a, Alhadeff & Freeze 1977.…”