Serratia plymuthica strain IC1270 isolated from the rhizosphere, possessing antagonistic activity towards a wide range of plant-pathogenic fungi, is able to hydrolyze phytate. Phytase activity was found intracellularly, while no activity was detected in the culture liquid. Optimum activity was found at pH 4-5; it completely disappeared at pH > 7.0 and 2.5. Phytase production was practically absent in the exponential phase and reached a maximum in the late stationary phase. Mutations of genes grrA and grrS, encoding GacA/GacS-like 2-component global regulatory system, or in gene rpoS encoding the sigma factor RpoS subunit of RNA polymerase, led to a deficiency in phytase production. Introduction into mutants of the respective wild-type genes cloned into the wide-range plasmid pJFF224-NX under the control of the bacteriophage T4 gene 32 promoter complemented this deficiency. This is the first report implicating the GacA/GacS global regulators and RpoS factor in phytase production in bacteria.

• MeSH
• 6-Phytase chemistry   genetics   metabolism   MeSH
• Bacterial Proteins chemistry   genetics   metabolism   MeSH
• Plant Roots microbiology   MeSH
• Gene Expression Regulation, Enzymologic * MeSH
• Gene Expression Regulation, Bacterial * MeSH
• Serratia chemistry   enzymology   genetics   MeSH
• Sigma Factor genetics   metabolism   MeSH
• Enzyme Stability MeSH
• Vitis microbiology   MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 6-Phytase MeSH
• Bacterial Proteins MeSH
• GacA protein, Bacteria MeSH Browser
• Sigma Factor MeSH

Cryptococcus laurentii strain AL27 demonstrating significant potential for intracellular phytase production was selected by 2-step screening of Antarctic yeasts. The strain showed increased phytase activity in a culture medium with 40 g/L sucrose, KH2PO4 providing 5 mg/L phosphorus, and cultivation temperature of 24 degrees C, which relates it to psychrotrophic microorganisms. The enzyme kinetic characteristics according to sodium phytate were Km = 0.98 mmol/L, vlim = 33.3 micromol g(-1) min(-1). The enzyme had maximum activity at 40 degrees C and acted within a wide pH range: from 2.0 to 5.5, which is of positive significance for its direct inclusion into the feed of monogastric animals.

• MeSH
• 6-Phytase chemistry   genetics   metabolism   MeSH
• Cryptococcus enzymology   genetics   isolation & purification   metabolism   MeSH
• Fungal Proteins chemistry   genetics   metabolism   MeSH
• Kinetics MeSH
• Soil Microbiology * MeSH
• Enzyme Stability MeSH
• Temperature MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Antarctic Regions MeSH
• Names of Substances
• 6-Phytase MeSH
• Fungal Proteins MeSH

The biochemical properties of the alkaline phosphatases (AlPs) produced by Rhizopus microsporus are described. High enzymic levels were produced within 1-2 d in agitated cultures with 1 % wheat bran. Intra- and extracellular AlPs were purified 5.0 and 9.3x, respectively, by DEAE-cellulose and ConA-sepharose chromatography. Molar mass of 118 and 120 kDa was estimated by gel filtration for both forms of phosphatases. SDS-PAGE indicated dimeric structures of 57 kDa for both forms. Mn(2+), Na(+) and Mg(2+) stimulated the activity, while Al(3+) and Zn(2+) activated only the extracellular form. Optimum temperature and pH for both phosphatases were 65 degrees C and pH 8.0, respectively. The enzymes were stable at 50 degrees C for at least 15 min. Hydrolysis of 4-nitrophenyl phosphate exhibited a K (m) 0.28 and 0.22 mmol/L, with upsilon (lim) 5.89 and 4.84 U/mg, for intra- and extracellular phosphatases, respectively. The properties of the reported AlPs may be suitable for biotechnological application.

• MeSH
• Alkaline Phosphatase isolation & purification   metabolism   MeSH
• Chromatography, Agarose MeSH
• Chromatography, DEAE-Cellulose MeSH
• Electrophoresis, Polyacrylamide Gel MeSH
• Fungal Proteins isolation & purification   metabolism   MeSH
• Chromatography, Gel MeSH
• Cations pharmacology   MeSH
• Hydrogen-Ion Concentration MeSH
• Molecular Weight MeSH
• Rhizopus enzymology   MeSH
• Hot Temperature MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Alkaline Phosphatase MeSH
• Fungal Proteins MeSH
• Cations MeSH

Biochemical properties of a termostable alkaline phosphatase obtained from the mycelium extract of A. caespitosus were described. The enzyme was purified 42-fold with 32% recovery by DEAE-cellulose and concanavalin A-Sepharose chromatography. The molar mass estimated by Sephacryl S-200 or by 7% SDS-PAGE was 138 kDa and 71 kDa, respectively, indicating a homodimer. Temperature and pH optima were 80 degrees C and pH 9.0. This enzyme was highly glycosylated (approximately 74% saccharide content). The activity was enhanced by Mg2+ (19-139%), NH4+ (64%), Na+ (51%) and Mn2+ (38%). 4-Nitrophenyl phosphate (4-NPP) was preferentially hydrolyzed, but glucose 1-phosphate (93%), UTP (67%) and O-phosphoamino acids also acted as substrates. V(lim) and K(m) were 3.78 nkat per mg protein and 270 micromol/L in the absence of Mg2+ and 7.35 nkat per mg protein and 410 micromol/L in the presence of Mg2+, using 4-NPP as substrate. The purified alkaline phosphatase removed the 5'-phosphate group of a linearized plasmid without showing DNAase activity, indicating its potential for recombinant DNA technology.

• MeSH
• Alkaline Phosphatase chemistry   isolation & purification   metabolism   MeSH
• Aspergillus enzymology   MeSH
• Deoxyribonucleases metabolism   MeSH
• Kinetics MeSH
• Mycelium enzymology   MeSH
• Substrate Specificity MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Alkaline Phosphatase MeSH
• Deoxyribonucleases MeSH

The production of phytase and associated feed enzymes (phosphatase, xylanase, CMCase, alpha-amylase and beta-glucosidase) was determined in a thermotolerant fungus Mucor indicus MTCC 6333, isolated from composting soil. Solid-substrate culturing on wheat bran and optimizing other culture conditions (C and N sources, level of N, temperature, pH, culture age, inoculum level), increased the yield of phytase from 266 +/- 0.2 to 513 +/- 0.4 nkat/g substrate dry mass. The culture extract also contained 112, 194, 171, 396, and 333 nkat/g substrate of phosphatase, xylanase, CMCase, beta-glucosidase and alpha-amylase activities, respectively. Simple 2-step purification employing anion exchange and gel filtration chromatography resulted in 21.9-fold purified phytase. The optimum pH and temperature were pH 6.0 and 70 degrees C, respectively. The phytase was thermostable under acidic conditions, showing 82% residual activity after exposure to 60 degrees C at pH 3.0 and 5.0 for 2 h, and displayed broad substrate specificity. The Km was 200 nmol/L and v(lim) of 113 nmol/s per mg protein with dodecasodium phytate as substrate. In vitro feed trial with feed enzyme resulted in the release of 1.68 g inorganic P/kg of feed after 6 h of incubation at 37 degrees C.

• MeSH
• 6-Phytase chemistry   isolation & purification   metabolism   MeSH
• alpha-Amylases metabolism   MeSH
• Cell Wall metabolism   MeSH
• Cellulase metabolism   MeSH
• Endo-1,4-beta Xylanases metabolism   MeSH
• Fermentation MeSH
• Chromatography, Gel MeSH
• Hydrolases metabolism   MeSH
• Hydrogen-Ion Concentration MeSH
• Animal Feed MeSH
• Culture Media chemistry   MeSH
• Mucor enzymology   growth & development   isolation & purification   MeSH
• Dietary Fiber microbiology   MeSH
• Industrial Microbiology * MeSH
• Enzyme Stability MeSH
• Substrate Specificity MeSH
• Temperature MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• 6-Phytase MeSH
• alpha-Amylases MeSH
• carboxymethylcellulase MeSH Browser
• Cellulase MeSH
• Endo-1,4-beta Xylanases MeSH
• Hydrolases MeSH
• Culture Media MeSH
• Dietary Fiber MeSH