Carbon, nitrogen and phosphorus mineralization in two soils amended with distillery yeast.

The possibility of using yeast from alcohol distilleries as a source of nutrients in soil was investigated. The following treatments were used: no fertilization (control), 0.5% (w/w) yeast, 1% (w/w) yeast, and NPK. The decomposition of yeast was monitored for 90 days in two soils. The CO2 production and the microbial biomass were increased by an average of 1- to 3-fold by yeast incorporation compared to control. Protease activity also was enhanced 3- to 8-fold in the soils supplemented with yeast compared to control. The phosphatase activities were higher than control only during the first days. While nitrate contents increased in all treatments compared to control, available P only increased in the soils amended with 1% yeast or NPK by 45-119% and 309-489%, respectively. These results indicate that there exists an excellent potential for the use of yeast in the soil as a source of nitrate and available P for plant nutrition.

Inorganic pyrophosphate-phosphohydrolytic activity associated with rat osseous plate alkaline phosphatase.

Purified membrane-bound alkaline phosphatase from rat osseous plate hydrolyzed pyrophosphate in the presence of magnesium ions, with a specific activity of 92.7 U/mg. Optimal apparent pH for pyrophosphatase activity was 8.0 and it remained unchanged on increasing the pyrophosphate concentration. In the absence of magnesium ions the enzyme had a Km = 88 microM and V = 36.7 U/mg for pyrophosphate and no inhibition by excess substrate was observed. Pyrophosphatase activity was rapidly destroyed at temperatures above 40 degrees C, but magnesium ions apparently protected the enzyme against denaturation. Sodium metavanadate (Ki = 1.0 mM) was a competitive inhibitor of pyrophosphatase activity, while levamisole (Ki = 8.2 mM) and theophylline (Ki = 7.4 mM) were uncompetitive inhibitors. Magnesium ions (K0.5 = 1.7 microM) stimulated pyrophosphatase activity, while cobalt (Ki = 48.5 microM) and zinc (Ki = 22.0 microM) ions were non-competitive inhibitors. Manganese and calcium ions had no effect on pyrophosphatase activity. The Mw of the pyrophosphatase protein was 130 kDa by gel filtration, but a value of 65 kDa was obtained by dissociative gel electrophoresis, suggesting that it was a dimer of apparently identical subunits. These results suggested that pyrophosphatase activity stems from the membrane-bound osseous plate alkaline phosphatase and not from a different protein.

Allosteric modulation of pyrophosphatase activity of rat osseous plate alkaline phosphatase by magnesium ions.

Pyrophosphatase activity of rat osseous plate alkaline phosphatase was studied at different concentrations of calcium and magnesium ions, with the aim of characterizing the modulation of enzyme activity by these metals. In the absence of metal ions, the enzyme hydrolysed pyrophosphate following "Michaelian" kinetics with a specific activity of 36.7 U/mg and K0.5 = 88 microM. In the presence of low concentrations (0.1 mM) of magnesium (or calcium) ions, the enzyme also exhibited "Michaelian" kinetics for the hydrolysis of pyrophosphate, but a significant increase in specific activity (123 U/mg) was observed, K(m) values remained almost unchanged. Quite different behavior occurred in the presence of 2 mM magnesium (or calcium) ions. In addition to low-affinity sites (K0.5-40 and 90 microM, for magnesium and calcium, respectively), high-affinity sites were also observed with K0.5 values 100-fold lower. The high-affinity sites observed in the presence of calcium ions represented about 10% of those observed for magnesium ions. This was correlated with the fact that only magnesium ions triggered conformational changes yielding a fully active enzyme. These results suggested that the enzyme could hydrolyse pyrophosphate, even at physiological concentrations (4 microM), since magnesium concentrations are high enough to trigger conformational changes increasing the enzyme activity. A model, suggesting the involvement of magnesium ions in the hydrolysis of pyrophosphate by rat osseous plate alkaline phosphatase is proposed.

Solubilization of membrane-bound matrix-induced alkaline phosphatase with polyoxyethylene 9-lauryl ether (polidocanol): purification and metalloenzyme properties.

1. Matrix-induced alkaline phosphatase prepared from rat osseous plate was solubilized with polidocanol and purified on a Sephacryl S-300 column. 2. Purified solubilized alkaline phosphatase has a molecular weight of ca 115,000 and bind one magnesium and two zinc ions. At least 110 detergent molecules are bound to each enzyme molecule. 3. Solubilization and purification procedures did not destroy the ability of the enzyme to hydrolyze adenosine-5'-triphosphate, p-nitrophenylphosphate, pyrophosphate and bis p-nitrophenylphosphate. 4. Magnesium, manganese and cobalt ions are stimulators of PNPPase activity of solubilized enzyme whereas calcium and zinc ions are inhibitors.