N-acetylglucosamine binding activity in extracts of adult newt skin.

1. Extracts of the dermis of the adult newt, Notophthalmus viridescens, contain hemagglutination activity which is specifically inhibited by N-acetylglucosamine. 2. The activity is soluble and is associated with a doublet in sodium dodecyl sulfate polyacrylamide gels, the bands of which have relative molecular weights of 51,000 and 57,000 under both reducing and non-reducing conditions. 3. The activity requires magnesium but not calcium, cobalt, or manganese and is inhibited by barium. 4. The activity is also dependent on pH with a pH optimum between 7.0 and 7.6.

Studies on an antineoplastic fraction from human urine. Characterization of the major protein in this fraction.

A fraction has been isolated from human urine which exhibits antiproliferative activity against human tumour cell lines without affecting the growth of several normal diploid cell lines or tumour cells of mouse or hamster origin. The major protein present in this fraction has been characterized and tentatively designated antineoplastic urinary protein (ANUP). An S020,W value of 3.69 S was obtained by sedimentation velocity analysis, and a subunit molecular mass of 16 300 Da was obtained by sedimentation equilibrium and by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Centrifugation data also indicated that the protein self-associates. The amino acid analysis of ANUP was consistent with its low pI (4.2) as determined by chromatofocusing analysis. Furthermore, the amino acid composition exhibited some features similar to collagen, as shown by high levels of proline and glycine, the absence of cysteine, and the presence of low levels of hydroxyproline.

Bovine milk xanthine oxidase: purification by ultrafiltration and conventional methods which omit addition of proteases: some criteria for homogeneity of native xanthine oxidase.

Methodological difficulties have been encountered when proteases were omitted from the conventional isolation of bovine milk xanthine oxidase (xanthine:oxygen oxidoreductase, EC 1.2.3.2). The use of these conventional methods has been studied and modified to reduce the problems encountered. Some of the difficulties may be due to the presence of high concentrations of caseins, which exhibit a wide range of charges and sizes, thereby making separations based on charge and size more complicated. In addition, non-covalent interactions may occur between the caseins and xanthine oxidase leading to the formation of casein-xanthine oxidase micellar aggregates. The difficulties encountered in this conventional isolation have been circumvented by purifying the enzyme directly from milk fat globule membranes that first have been washed free of most casein and other milk proteins. The xanthine oxidase is isolated by ultrafiltration through an Amicon XM-100A membrane at 5 degrees C in 0.25 M sucrose/5 mM sodium salicylate. The largest molecular size of globular proteins which can penetrate this ultrafiltration membrane has been previously estimated to be around 100 000 daltons. Xanthine oxidase thus appears to be smaller than 100 000 daltons in its native state. The size observed for active xanthine oxidase previously isolated by other methods has been around 275 000--300 000 daltons. Xanthine oxidase isolated by ultrafiltration appears similar to xanthine oxidase from conventional isolation methods according to empirical criteria of homogeneity based on size and also on the absorbances at 280 and 450 nm. Criteria based on charge were found to be less reliable.