N-butanol Treatment Research Articles

Studies on lysophospholipases: IV. The subcellular distribution of two lysolecithin-hydrolyzing enzymes in beef liver

1. 1. In a previous paper (Biochim. Biophys. Acta (1974) 369, 50–63) the purification of two proteins with lysophospholipase activity (EC 3.1.1.5), provisionally denoted lysophospholipase I and lysophospholipase II, has been described. The subcellular localization of both enzymes was investigated by cell fractionation studies. 2. 2. For each subcellular fraction the total lysophospholipase activity, after solubilization by n-butanol treatment, was separated into a lysophospholipase I and II contribution by DEAE-Sephadex ion exchange chromatography. 3. 3. Lysophospholipase I was found to be a soluble enzyme with a bimodal distribution. Highest relative specific activities were measured in the mitochondrial and the cytoplasmic fraction. Evidence is presented indicating that this enzyme is present in the mitochondrial matrix fraction. 4. 4. Lysophospholipase II appeared to be a membrane-bound enzyme with highest relative specific activity in the microsomal fraction.

Purification and characterization of hexose oxidase from the red alga Chondrus crispus

Hexose oxidase ( d-hexose:O 2 oxidoreductase, EC 1.1.3.5) from the red alga Chondrus crispus has been purified by a procedure involving extraction with 0.1 M sodium phosphate (pH 6.8), n-butanol treatment, (NH 4) 2SO 4 precipitation, DEAE-cellulose chromatography, pepsin-trypsin digestion, and gel filtration on Sephadex G-200. The purified enzyme shows only a single band on disc gel electrophoresis. The enzyme has a mol. wt of 130 000 as determined by gel filtration and contains approximately 12 gram-atoms of copper per mole. FAD was not detected in the enzyme. The enzyme is a glycoprotein containing about 70% of a carbohydrate moiety consisting mainly of galactose and xylose. Optimum temperature and pH of the enzyme are 25 °C and 6.3, respectively. The enzyme oxidizes d-glucose, d-galactose, maltose, cellobiose, and lactose. The products of hexose oxidation are H 2O 2 and hexonolactone. The enzyme is strongly inhibited by diethyldithiocarbamate and to a less extent by cyanide, azide, hydroxylamine, and acetate. The production of H 2O 2 by the enzyme is responsible for the toxicity of Chondrus crispus to Chlorella. The red alga Euthora cristata also appears to contain this enzyme.

An improved resolution of cytoplasmic proteins of rat liver after n-butanol treatment.

IN an earlier communication1, a technique was described for obtaining consistent separations of the cytoplasmic proteins of rat liver using electrophoresis on filter paper. A notable improvement in resolution has been observed following the treatment of the protein mixtures with n-butanol. The use of this reagent was suggested by the experiments of Morton2 on the extraction of various enzymes from the particulate fractions of the cell structure.