02656nas a2200253   4500008004100000022001400041245014200055210006900197260009900266300001400365490000700379520163400386653002402020653004402044653001702088653002402105653002402129100001702153700002602170700002002196700003002216700002602246856013002272       2006                        eng d  a1359-511300aPurification, characterization and substrate specificity of thermostable alpha-galactosidase from Bacillus stearothermophilus (NCIM-5146)0 aPurification characterization and substrate specificity of therm  aTHE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, OXON, ENGLANDbELSEVIER SCI LTDcJUN  a1311-13170 v413 a<p>An extracellular thermostable alpha-galactosidase from Bacillus stearothermophilus (NCIM-5146) has been purified to homogeneity by chromatographic step, using Phenyl Sepharose CL-4B column. The specific activity of the enzyme was increased approximately 389-fold, from 1.03 U/mg protein to 400 U/mg protein. The molecular mass of the purified enzyme as determined by SDS-PAGE and gel filtration was 79.9 and 165.9 kDa, respectively, suggesting dimeric nature. The purified alpha-galactosidase is a non-glycosylated protein with a pI of 4.9. The pH and temperature optima for the purified enzyme are 6.5-7.0 and 65 degrees C, respectively. The alpha-galactosidase is stable over a broad pH range (3-9) and its half-life of inactivation (t(1/2)) at 70 degrees C is 30 min. The partial N-terminal sequence of alpha-galactosidase showed remarkable homology (80% similarity) with earlier reported alpha-galactosidase from B. stearothermophilus NUB 3621. The secondary structure of the enzyme determined by circular dichroism (CD) spectroscopy exhibited alpha/beta class of protein and showed temperature induced conformational forms below and above the transition temperature. The purified enzyme showed biphasic Arrhenius plot with break point at 55 degrees C for pNPG and 50 degrees C for melibiose, raffinose and stachyose. The enzyme hydrolyzes alpha-1-3, alpha-1-4, and alpha-1-6 galactosidic linkages and not the beta-galactosidic linkages. Synthetic substrates pNPG and oNPG had lower K-m and higher K-cat as compare to natural substrates, melibiose, raffinose, and stachyose. (c) 2006 Elsevier Ltd. All rights reserved.</p>10aalpha-galactosidase10aBacillus stearothermophilus (NCIM-5146)10aPurification10aSecondary structure10athermostable enzyme1 aGote, M., M.1 aKhan, Mohammad, Islam1 aGokhale, D., V.1 aBastawade, Kulbhushan, B.1 aKhire, Jayant, Malhar  uhttps://library.ncl.res.in/content/purification-characterization-and-substrate-specificity-thermostable-alpha-galactosidase-002418nas a2200229   4500008004100000022001400041245012600055210006900181260005700250300001400307490000700321520159900328653002201927653001201949653001201961653001701973653001701990100001702007700001502024700002602039856012302065       2010                        eng d  a0959-399300aPurification and characterization of two distinct acidic phytases with broad pH stability from aspergillus niger NCIM 5630 aPurification and characterization of two distinct acidic phytase  a233 SPRING ST, NEW YORK, NY 10013 USAbSPRINGERcNOV  a2009-20180 v263 a<p>Aspergillus niger NCIM 563 produced two different extracellular phytases (Phy I and Phy II) under submerged fermentation conditions at 30A degrees C in medium containing dextrin-glucose-sodium nitrate-salts. Both the enzymes were purified to homogeneity using Rotavapor concentration, Phenyl-Sepharose column chromatography and Sephacryl S-200 gel filtration. The molecular mass of Phy I and II as determined by SDS-PAGE and gel filtration were 66, 264, 150 and 148 kDa respectively, indicating that Phy I consists of four identical subunits and Phy II is a monomer. The pI values of Phy I and II were 3.55 and 3.91, respectively. Phy I was highly acidic with optimum pH of 2.5 and was stable over a broad pH range (1.5-9.0) while Phy II showed a pH optimum of 5.0 with stability in the range of pH 3.5-9.0. Phy I exhibited very broad substrate specificity while Phy II was more specific for sodium phytate. Similarly Phy II was strongly inhibited by Ag(+), Hg(2+) (1 mM) metal ions and Phy I was partially inhibited. Peptide analysis by Mass Spectrometry (MS) MALDI-TOF also indicated that both the proteins were totally different. The K (m) for Phy I and II for sodium phytate was 2.01 and 0.145 mM while V (max) was 5,018 and 1,671 mu mol min(-1) mg(-1), respectively. The N-terminal amino acid sequences of Phy I and Phy II were FSYGAAIPQQ and GVDERFPYTG, respectively. Phy II showed no homology with Phy I and any other known phytases from the literature suggesting its unique nature. This, according to us, is the first report of two distinct novel phytases from Aspergillus niger.</p>10aAspergillus niger10aPhytase10aPhytate10aPoultry feed10aPurification1 aSoni, S., K.1 aMagdum, A.1 aKhire, Jayant, Malhar  uhttps://library.ncl.res.in/content/purification-and-characterization-two-distinct-acidic-phytases-broad-ph-stability-002146nas a2200229   4500008004100000022001400041245018200055210006900237260009900306300001400405490000700419520119100426653002201617653002901639653001201668653002401680653001701704100002401721700001601745700002601761856012901787       2012                        eng d  a1359-511300aDownstream processing of extracellular phytase from aspergillus niger: chromatography process vs. aqueous two phase extraction for its simultaneous partitioning and purification0 aDownstream processing of extracellular phytase from aspergillus   aTHE BOULEVARD, LANGFORD LANE, KIDLINGTON, OXFORD OX5 1GB, OXON, ENGLANDbELSEVIER SCI LTDcJUL  a1066-10720 v473 a<p>The application of single step aqueous two-phase extraction (ATPE) for the downstream processing of phytase from Aspergillus niger NCIM 563, produced under solid state fermentation, has been studied and compared with the traditional multi-step procedure involving salt precipitation and column chromatography. High phytase recovery (98.5%) within a short time (3 h) and improved thermostability was attained by ATPE in comparison to 20% recovery in 96 h by chromatography process.The ATPE method, therefore, seems to be an interesting alternative for simultaneous partitioning and purification of phytase. The influence of system parameters; such as, phase forming salts, polymer molecular weight and system pH on the partitioning behavior of phytase was evaluated. The ATPE system consisting of combination of polyethylene glycol (PEG) 6000 and 8000(10.5%) and sodium citrate (20.5%) resulted in one-sided partitioning of phytase in bottom phase with a purification factor of 2.5. This is the first report on phytase purification using liquid-liquid extraction and the results are likely to be beneficial in the poultry feed industry. (c) 2012 Elsevier Ltd. All rights reserved.</p>10aAspergillus niger10aliquid-liquid extraction10aPhytase10aPolyethylene glycol10aPurification1 aBhavsar, Kavita, P.1 aKumar, Ravi1 aKhire, Jayant, Malhar  uhttps://library.ncl.res.in/content/downstream-processing-extracellular-phytase-aspergillus-niger-chromatography-process-vs-0