Nitrogenase activity in extracts of heterocystous and non-heterocystous blue-green algae

Summary Cell-free extracts capable of acetylene reduction and cyanide reduction have been prepared from heterocystous (Anabaena cylindrica) and non-heterocystous (Plectonema boryanum 594) blue-green algae. Extracts from Anabaena were obtained from cultures grown in blulk under aerobic conditions, while the Plectonema cultures were grown in bulk on nitrate-nitrogen, then washed free from nitrate and sparged with A/CO₂ for 40 h after which time maximum nitrogenase activity was detected. The nitrogenases of both algae are similar and resemble in many respects nitrogenases from bacteria and legumes. Activity is located primarily in a 40,000xgx15 min supernatant fraction and the rate of C₂H₂ reduction observed is about 10 per cent of whole cell activity. ATP and a source of reducing power (Na₂S₂O₄) are required for efficient functioning of the enzyme. ATP-dependent hydrogen evolution occurs, the extracts are cold labile and highly sensitive to oxygen and the oxygen inhibition is irreversible.     

ANTIBODY-PRODUCING CELLS: ANALYSIS AND PURIFICATION BY VELOCITY SEDIMENTATION

Velocity sedimentation cell separation is a simple and reproducible method for obtaining highly enriched populations of viable antibody-producing cells. Using suspensions of spleen cells prepared from mice immunized with sheep erythrocytes, fractions containing up to 2% 19S-PFC and 25% 7S-PFC can be obtained. Granulocytes constitute almost all of the remaining cells in these fractions. The sedimentation profile of 7S-PFC is very broad in comparison with that of cell populations known to be homogeneous in size (e.g. mouse erythrocytes). Analysis of the profile of 7S-PFC at different times after immunization suggests that the heterogeneity arises largely from the doubling in cell volume as a cell moves from one mitosis to the next. Early in the immune response, when the majority of the PFC are proliferating, the variation in sedimentation velocities is consistent with such a two-fold variation in cell volume. Late in the response, when most PFC have stopped proliferating, the sedimentation profile is more homogeneous. This analysis suggests that the fractionation procedure is sensitive enough to separate PFC according to their position in the cell cycle. Sedimentation velocities were also measured for several other classes of cells found in spleen. Comparison of these values shows that sedimentation velocity is a useful parameter for characterizing different types of cells.     

Lipolytic Esterases in Staphylococci

Staphylococci split a wide range of lipid substrates by production of an enzyme complex with two main components (i) a lipase acting optimally on fat-soluble glycerides, and (ii) an esterase acting optimally on water-soluble esters. The action is dependent upon carbon chain length, interfacial dispersion, solubility, and pH of substrate and end products. The esterase is less susceptible to organophosphorus inhibitors than mammalian esterases. There is no apparent correlation between lipolysis and markers of pathogenicity such as production of coagulase and toxin, but the possession of a flexible lipolytic mechanism might account for the persistence of staphylococci in the fatty secretions of mammalian skin.