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1Author    K. Irumakki, N. ShivaramRequires cookie*
 Title    Purification and Properties of Potato Phosphorylase Isozymes  
 Abstract    Two multiple forms of a-glucan phosphorylase which migrate about half way in polyacryl-amide-gel electrophoresis (named "slow" and "fast" isozyme), were isolated by combined chromato­ graphy and preparative electrophoresis after freezing the tissue from freshly harvested and from sprouting potato tubers respectively. Depending on the primer used for the synthesis reaction their pH optimum varied between 5.2 and 6.0 and the optimum temperature was 30 and 35 °C. The isoelectric point for the slow isozyme was at pH 5 .0 + 0.1 and for the fast isozyme, pH 5 .5 + 0.1, the molecular weights were 209 000 + 10 000 and 165 000 + 5000 and for their subunits 104 000 + 4000 and 40 000 + 2000 respectively. Both isozymes were inhibited by Hg2+, Ag+ and p-chloromercuro-benzoate (p-CMB). Fe2+ ions inhibited them partially. Mg2+, Mn2+ and sulfhydryl compounds acti­ vated both. K m values for the slow and fast isozymes with glucose-l-phosphate in presence of soluble starch was 6.7 and 8.0 mM, of amylose 14.3 and 20.0 mM and of glycogen 22.2 and 40.0 mM respectively. The affinity of the primer for the slow and the fast isozymes were as follows: soluble starch 0.5 and 1.0 mM, amylose 2.6 and 3.8 mM, glycogen 6.2 and 7.7 mM respectively. K m values of phos-phorolysis with soluble starch was 0.8 and 0.5 mM, with amylose was 3.1 and 1.1 mM, and with glycogen was 6.5 and 1.3 mM respectively. As substrate and primer the soluble starch was superior and the glycogen was inferior. Amylose was in between. Kinetic parameters suggested the existence of a-glucan phosphorylase isozymes with different specificities: the slow one being more active in the direction of starch synthesis and the fast iso­ zyme degrading faster the polyglucans. These observations suggest that the polyglucan synthesis and degradation in potato tubers may be regulated by the change in the proportion of slow and fast isozymes. ' 
  Reference    (Z. Naturforsch. 31c, 424 [1976]; received April 5 1976) 
  Published    1976 
  Keywords    Potato, Phosphorylase, Isozymes, Purification, Properties 
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 TEI-XML for    default:Reihe_C/31/ZNC-1976-31c-0424.pdf 
 Identifier    ZNC-1976-31c-0424 
 Volume    31 
2Author    Oskar Oster, Gerhard BuchlowRequires cookie*
 Title    Purification of Histone F3 by Covalent Chromatography  
 Abstract    The arginine-rich histone F3 has been purified by covalent affinity chromatography. By the use of activated Thiol-Sepharose 4B for the purification of cysteine containing histone F3 a highly pure protein was obtained. The simple purification procedure offers the opportunity to get larger amounts of pure histone F3 within short time. 
  Reference    (Z. Naturforsch. 32c, 72 [1977]; received October 18 1976) 
  Published    1977 
  Keywords    Histone F3, Purification, Covalent Chromatography 
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 TEI-XML for    default:Reihe_C/32/ZNC-1977-32c-0072.pdf 
 Identifier    ZNC-1977-32c-0072 
 Volume    32 
3Author    Ingrid Gentzen, Hans-G Löffler, Friedh SchneiderRequires cookie*
 Title    Aminoacylase from Aspergillus oryzae. Comparison with the Pig Kidney Enzyme  
 Abstract    Am inoacylase (EC 3.5.1.14) from Aspergillus oryzae was purified from a com m ercially available crude material by heat treatment, precipitation by polyethyleneim ine and amm onium sulfate, gel chromatography and preparative disc-gel-electrophoresis. The purified product was hom ogenous as judged by polyacrylamide gel electrophoresis. SDS-gel electrophoresis, polyacrylam ide-gel-gradient electrophoresis, gel chromatography and amino acid analysis demonstrated the enzyme to be com posed o f two subunits with M r o f 36 600. The kinetic properties o f the enzyme were studied with chloroacetyl derivatives o f alanine, phenylalanine, methionine, leucine, norleucine and tryptophan. The pH optim um o f the acylase activity with chloroacetyl-alanine as substrate is at pH 8.5. Acyl derivatives o f hydrophobic am ino acids are preferred substrates. The enzyme has no dipeptidase activity. Am inoacylase is not inhibited by SH -blocking agents and no SH-groups could be detected with Ellman's reagent in the native and denatured enzyme. The enzyme activity is insensitive to phenylmethylsulfonyl fluoride and N-a-/?-tosyl-L-lysine chlorom ethyl ketone. The microbial acylase is a zinc m etallo enzyme. M etal chelating agents are strong inhibitors; it is further inhibited by Cd2+, Mn2+, N i2+, C u2+ and activated by C o2+. The properties o f pig kidney and Aspergillus acylase are compared. 
  Reference    Z. Naturforsch. 35c, 544 (1980); received February 21 1980 
  Published    1980 
  Keywords    Aspergillus Am inoacylase, Purification, Properties 
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 TEI-XML for    default:Reihe_C/35/ZNC-1980-35c-0544.pdf 
 Identifier    ZNC-1980-35c-0544 
 Volume    35 
4Author    Cornelia Fuchs, G.Erd HansenRequires cookie*
 Title    Partial Purification and Some Properties of Brassica napus Lipase  
 Abstract    Lipase (triacylglycerol acylhydrolase EC 3.1.1.3) from rape (Brassica napus cv. Ceres) was isolated from cotyledons of dark-grown seedlings. The enzyme was partially purified by poly­ ethylene glycol precipitation. Delipidation of the lipase with «-hexane was required prior to further purification by anion exchange chromatography and size exclusion chromatography. A purification factor of 337 was ultimately achieved and the purification process was moni­ tored by SDS-PAGE. Here, at least two protein bands with molecular masses of 62 and 64 kD a respectively were found in the active fraction obtained by size exclusion chrom atogra­ phy. Sodium deoxycholate was found to stimulate the lipase activity, but appeared to cause aggregation of the enzyme. It was not possible to estimate the isoelectric point of the dialyzed rape lipase due to the high molecular mass of the aggregates. Two simple methods to detect lipase activity directly on polyacrylamide gel were applied. No esterase activity was found by using p-nitrophenyl acetate as substrate. 
  Reference    Z. Naturforsch. 49c, 293—301 (1994); received November 3 1993/March 7 1994 
  Published    1994 
  Keywords    Lipase, Rape, Purification, Properties 
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 TEI-XML for    default:Reihe_C/49/ZNC-1994-49c-0293.pdf 
 Identifier    ZNC-1994-49c-0293 
 Volume    49 
5Author    Toshihisa Ohshima, Gerhart DrewsRequires cookie*
 Title    Isolation and Partial Characterization of the Membrane-Bound NADH Dehydrogenase from the Phototrophic Bacterium Rhodopseudomonas capsulata  
 Abstract    Chem otrophically grown cells of Rhodopseudomonas capsulata contain at least three different pyridine nucleotide dehydrogenases, i) a soluble, found in the supernatant (144000 x g) o f cell free extracts, N AD H-dependent, ii) a mem brane-bound, N AD H -dependent, and iii) a soluble, found in the supernatant N AD PH dependent. i The m em brane-bound N A D H dehydrogenase (E.C. 1.6.99.3) has been solubilized by sodium deoxycholate treatm ent of m em branes and purified 75 fold by column chrom atography on Sephadex G-150 and DEAE cellulose in the presence of sodium cholate. The native enzyme has an apparent molecular mass (M r) o f 97 000, containing polypeptides of Mr of about 15 000. The pH optim um was at 7.5. The enzyme was specific for NADH. The Michaelis constant for NADH and DCIP were 4.0 and 63 hm, respectively. The enzyme was inactivated by FM N, riboflavin and NADH. In contrast, the soluble N ADH-dehydrogenase (i) was activated by FMN. 
  Reference    Z. Naturforsch. 36c, 400 (1981); received February 23/M arch 16 1981 
  Published    1981 
  Keywords    NADH Dehydrogenase, Purification, Localization, Inhibitors, Rhodopseudomonas capsulata 
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 TEI-XML for    default:Reihe_C/36/ZNC-1981-36c-0400.pdf 
 Identifier    ZNC-1981-36c-0400 
 Volume    36 
6Author    Zsuzsa Izsvák, Zsolt Jobbágy, Ernö DudaRequires cookie*
 Title    Purification and Characterization of Ceql Restriction Endonuclease  
 Abstract    C eql, a type II restriction endonuclease, an isoschizomer of EcoRV was purified to apparent homogeneity by a combination of salt precipitation, ion exchange, dye affinity and hydrophobic interaction chromatographies. The crude enzyme was present in the form of large aggregates that could be pelleted by high speed centrifugation. The enzyme was not associated with cellular membranes, though non-ionic detergents lowered the apparent size of the aggregates. The puri­ fied enzyme also showed a tendency to form large molecular mass (66-600 kDa) complexes under physiological conditions, in the absence of cleavable DN A. The enzyme formed smaller complexes in the presence of D N A and non-ionic detergents and dissociated into subunits (and undergoes reversible loss of activity) in the presence of high concentrations of salts. According to SDS gel electrophoresis and sedimentation analysis the molecular mass of the monomer 32 ± 2 kDa. The enzyme had a rather broad pH optimum, extending into the alka­ line range and lost specificity and activity in buffers below pH 6. 
  Reference    Z. Naturforsch. 47c, 830—834 (1992); received May 26/ 
  Published    1992 
  Keywords    Restriction Enzyme, Endonuclease, Purification, Corynebacterium, Multimeric Enzyme 
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 TEI-XML for    default:Reihe_C/47/ZNC-1992-47c-0830.pdf 
 Identifier    ZNC-1992-47c-0830 
 Volume    47 
7Author    AniruddhaP. Sane, VidhuA. SaneRequires cookie*
 Title    Isolation and Kinetic Analyses of the Soluble ¥i ATPases from Mitochondria of Wheat and Pearlmillet  
 Abstract    The mitochondrial F j ATPases from two cereal crops, wheat and pearlmillet, were purified and studied. The wheat F, ATPase could be purified to homogeneity and is apparently com­ posed of six subunits with apparent molecular weights of 55 kDa (a and ß), 35 kDa (y), 26 kDa (8 ') and 22 kDa (8). The e subunit was barely detectable. Both enzymes reveal typical non-linear kinetics but show variability in their response to bicarbonate and chloride. While the wheat F, ATPase is stimulated by bicarbonate and chloride, the pearlmillet F j ATPase is inhibited by both anions. The two enzymes are Mg2+ dependent ATPases and are competi­ tively inhibited by Ca2+, unlike maize, pea and turnip ATPases. Both the enzymes also pos­ sess a GTPase activity which is two fold higher than the ATPase, unlike rice, sorghum and oat root Fj ATPases. 
  Reference    Z. Naturforsch. 53c, 341 (1998); received December 12 1997/January 22 1998 
  Published    1998 
  Keywords    F] ATPase, Purification, Kinetics, Wheat, Pearlmillet 
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 TEI-XML for    default:Reihe_C/53/ZNC-1998-53c-0341.pdf 
 Identifier    ZNC-1998-53c-0341 
 Volume    53 
8Author    M.Arianne Nagel, ThomasH. Artm AnnRequires cookie*
 Title    Glutamate Dehydrogenase from Medicago sativa L., Purification and Comparative Kinetic Studies of the Organ-Specific Multiple Forms  
 Abstract    NAD-specific glutamate dehydrogenase [L-glutamate: N A D + oxidoreductase (deaminating) EC 1.4.1.2] from Medicago sativa constitutes organ-specific patterns o f isoenzymes. The isoenzyme-pattems o f seeds (GDH-I) and roots (GDH-II) were purified 1520-fold and 92-fold, respectively. All isoenzymes o f both patterns remain stable throughout the purification procedures. Isoenzyme a7, the only isoenzyme common to both patterns was isolated from the GDH-I pattern. The three enzyme preparations were found to be identical in pH optima, substrate specificity and general ki­ netic properties. A comparative kinetic analysis revealed no pronounced differences between the various kinetic constants evaluated for the three enzyme preparations. Furthermore an identical order of substrate binding and product release could be established. Both initial rate measure­ ments and product inhibition studies are consistent with an ordered ternary-binary kinetic mecha­ nism. The results suggest that tissue-specific enzyme multiplicity of plant glutamate dehydrogen­ ase is not related to differences in general or kinetic properties. 
  Reference    Z. Naturforsch. 35c, 406 (1980); received January 71980 
  Published    1980 
  Keywords    Medicago sativa, Glutamate Dehydrogenase, Multiple Enzyme Forms, Purification, Kinetic Pro­ perties 
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 TEI-XML for    default:Reihe_C/35/ZNC-1980-35c-0406.pdf 
 Identifier    ZNC-1980-35c-0406 
 Volume    35 
9Author    Heinz-W Alter, Scheid, Adelheid Ehmke, Thomas HartmRequires cookie*
 Title    Plant NAD-Dependent Glutamate Dehydrogenase. Purification, Molecular Properties and Metal Ion Activation of the Enzymes from Lemna minor and Pisum sativum  
 Abstract    Glutamate dehydrogenase (L-glutamate: N A D + oxidoreductase (deaminating) EC 1.4.1.2) has been purified to homogeneity from Lemna minor and seeds o f Pisum sativum. As established by polyacrylamide gel electrophoresis the Pisum-enzyme constitutes a multiple pattern o f seven char­ ge isoenzymes whereas the Lemna enzyme shows one single protein band. Molecular weights of 230 000 were calculated for both enzymes by sedimentation equilibrium measurements (Pisum-enzyme]I and comparative gel filtration (Lemna-enzyme). Sodium dodecyl sulfate gel electropho­ resis and electron microscopic observations revealed that both enzymes are composed of four identical subunits (molecular weight 58 500) arranged in a tetraedric structure. The amino acid compositions of both enzymes are similar to those of various hexameric glutamate dehydrogen­ ases. The N-terminal amino acid of the Pisum-enzyme is alanine. Both enzymes require Ca2+ for maximal catalytic activity. For the Lemna-enzyme the K0.s values for Ca2+ are 22 /j.m (NADH-de-pendent reaction) and 4 piM (N A D +-dependent reaction), respectively. Ca2+ which to some extent can be replaced by Zn2+ does not affect the enzyme aggregation but seems to govern a reversible equilibrium between catalytically active and inactive enzyme forms. 
  Reference    Z. Naturforsch. 35c, 213—221 (1980); received November 21 1979 
  Published    1980 
  Keywords    Lemna minor, Pisum sativum, Glutamate Dehydrogenase, Purification, Molecular Properties, Me­ tal Ion Activation 
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 TEI-XML for    default:Reihe_C/35/ZNC-1980-35c-0213.pdf 
 Identifier    ZNC-1980-35c-0213 
 Volume    35 
10Author    Hendrik Hüdig, Gerhart DrewsRequires cookie*
 Title    Isolation of a b-Type Cytochrome Oxidase from Membranes of the Phototrophic Bacterium Rhodopseudomonas capsulata  
 Abstract    A cytochrome oxidase (EC 1.9.3.1) was solubilized from the membrane fraction o f aerobically grown cells of Rhodopseudomonas capsulata by treatment with Triton X-100. The enzyme was purified 160 fold by chromatography on DEAE-Sepharose CL -6B and affinity chromatography on cytochrome c-thiol activated Sepharose 4B. The purified enzyme has a pH-optimum at 8.5 and a temperature optimum at 35 °C. The ap­ parent K m for reduced horse cytochrome c is 24 |iM (at pH 8 and 30 °C). The purified cytochrome oxidase was 50% inhibited by 1.5 |iM KCN and 10 (iM N aN ,. The purified enzyme contained one polypeptide of mT 65,000 and 6-type cytochrome. 
  Reference    Z. Naturforsch. 37c, 193—198 (1982); received November 2 December 1 1981 
  Published    1982 
  Keywords    Rhodopseudomonas capsulata, Purification, Solubilization, Cytochrome c, Cytochrome Oxidase, 6-Type Cytochrome 
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 TEI-XML for    default:Reihe_C/37/ZNC-1982-37c-0193.pdf 
 Identifier    ZNC-1982-37c-0193 
 Volume    37