Stimulation of β-(l → 6)-glucanase production by oxidized pustulan

1972 ◽  
Vol 18 (10) ◽  
pp. 1543-1550 ◽  
Author(s):  
Robert G. Brown
Keyword(s):  
Enzyme Activity ◽  
Isoelectric Focusing ◽  
Gel Filtration ◽  
Tween 80 ◽  
Sole Source ◽  
Cellulase Production ◽  
Low Degree ◽  
Degree Of Oxidation ◽  
High Degree ◽  
Stimulation Of

A strain of Penicillium lilacinum, isolated from soil, produced pustulanase, β-(1 → 3)-glucanase, (EC. 3.2.1.6) and cellulase (EC.3.2.1.4) when cultivated on a medium containing pustulan as the sole source of carbon. If pustulan was replaced by ketopustulan, the production of pustulanase was stimulated about 10-fold although the amount of stimulation was dependent on the degree of oxidation of pustulan. β-(1 → 3)-Glucanase production was stimulated slightly by ketopustulan; however, the degree of oxidation did not affect significantly the yield of this enzyme. Cellulase production was either unaffected by the oxidized polymer, or at higher degrees of oxidation, decreased. Tween 80 stimulated the production of the three enzymes in media containing ketopustulan with a low degree of oxidation but was inhibitory to pustulanase and cellulase production in media containing ketopustulan with a high degree of oxidation. A combination of gel filtration and isoelectric focusing revealed that each enzyme activity was attributable to at least two proteins.

scholarly journals Cathepsin D. Purification of isoenzymes from human and chicken liver

1970 ◽  
Vol 117 (3) ◽  
pp. 601-607 ◽  
Author(s):  
A. J. Barrett
Keyword(s):  
Isoelectric Focusing ◽  
Cathepsin D ◽  
Ph Dependence ◽  
Gel Filtration ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Ph Optimum ◽  
Specific Activities ◽  
Acetone Fractionation ◽  
High Degree

1. The Barrett (1967) assay for cathepsin D was slightly modified. 2. The enzyme was purified from liver of man and chicken by a procedure involving autolysis, acetone fractionation, ion-exchange chromatography and isoelectric focusing. 3. Several isoenzymes of cathepsin D were resolved in the isoelectric-focusing step, and three major forms, α,β and γ, were distinguished for each species. 4. A modified analytical method of isoelectric focusing in polyacrylamide gel indicated a high degree of homogeneity of the purified β and γ isoenzymes from each species, and this was supported by their constant high specific activities. 5. Gel filtration of the isoenzymes in a calibrated column of Sephadex G-100 showed that each had a molecular weight of 45000. 6. Human cathepsin D had a pH optimum of 3.5, and that of chicken enzyme was 3.0, haemoglobin being used as substrate. In each species, the three isoenzymes have the same pH-dependence curve. 7. The purified cathepsin D samples showed very little action on acid-denatured albumin.

scholarly journals The Effects of Different Degrees of Procyanidin Polymerization on the Nutrient Absorption and Digestive Enzyme Activity in Mice

Molecules ◽  
2018 ◽  
Vol 23 (11) ◽  
pp. 2916 ◽  
Author(s):  
Huairong Zhong ◽  
Yong Xue ◽  
Xiaoyuan Lu ◽  
Qiang Shao ◽  
Yuelei Cao ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Amylase Activity ◽  
Digestive Enzyme ◽  
Degree Of Polymerization ◽  
Nutrient Absorption ◽  
Digestive Enzyme Activity ◽  
Total Food Intake ◽  
Total Food ◽  
Low Degree ◽  
High Degree

Proanthocyanidins, including polymers with both low and high degrees of polymerization, are the focus of intensive research worldwide due to their high antioxidant activity, medicinal applications, and pharmacological properties. However, the nutritional value of these compounds is limited because they readily form complexes with proteins, polysaccharides, and metal ions when consumed. In this study, we examined the effects of proanthocyanidins with different degrees of polymerization on white mice. Twenty-four male white mice were randomly divided into three groups of eight mice each and fed proanthocyanidins with a low degree of polymerization or a high degree of polymerization or a distilled water control via oral gavage over a 56-day period. We examined the effects of these proanthocyanidins on digestive enzyme activity and nutrient absorption. Compared to the control group, the group fed high-polymer proanthocyanidins exhibited a significant reduction in net body mass, total food intake, food utility rate, amylase activity, protease activity, and major nutrient digestibility (p < 0.05), while the group fed low-polymerization proanthocyanidins only exhibited significant reductions in total food intake, α-amylase activity, and apparent digestibility of calcium and zinc (p < 0.05). Therefore, proanthocyanidins with a high degree of polymerization had a greater effect on digestive enzyme activity and nutrient absorption than did those with a low degree of polymerization. This study lays the foundation for elucidating the relationship between procyanidin polymerization and nutrient uptake, with the aim of reducing or eliminating the antinutritional effects of polyphenols.

Trimethoprim resistance in Haemophilus influenzae is due to altered dihydrofolate reductase(s)

1991 ◽  
Vol 274 (3) ◽  
pp. 657-662 ◽  
Author(s):  
R de Groot ◽  
D O Chaffin ◽  
M Kuehn ◽  
A L Smith
Keyword(s):  
Enzyme Activity ◽  
Haemophilus Influenzae ◽  
Molecular Mass ◽  
Isoelectric Focusing ◽  
Dihydrofolate Reductase ◽  
Peptide Mapping ◽  
Gel Filtration ◽  
Peptide Fragment ◽  
Total Enzyme Activity ◽  
Total Enzyme

We characterized a highly purified preparation of the chromosomally encoded dihydrofolate reductase (DHFR) from a trimethoprim-susceptible (Tmp8; strain MAP) and two trimethoprim-resistant (TmpR) strains (MAP/47 and MAP/42) of Haemophilus influenzae. The enzymes were purified between 650- and 3000-fold by gel-filtration and dye-ligand chromatography. The apparent molecular mass of the three proteins was 18400 Da by PAGE under denaturing and nondenaturing conditions. Total enzyme activity was greater in all fractions from the TmpR strains compared with the Tmp8 isolate. The three enzymes had a similar Km for dihydrofolate (7, 9 and 5 microM) and NADPH (2, 5 and 6 microM). However, the Tmp IC50 (the concentration necessary for 50% inhibition of DHFR activity) for the Tmp8 strain MAP was 0.001 microM, whereas DHFR from the TmpR strains MAP/47 and MAP/42 had values of 0.1 microM and 0.3 microM respectively. The methotrexate IC50 of the MAP/42 DHFR was 0.06 microM in comparison with the enzyme from MAP (0.008 microM) and MAP/47 (0.007 microM). Isoelectric focusing indicated that the DHFR from MAP/42 had a different isoelectric point (pI 7.6) compared with the enzymes from MAP and MAP/47 (pI 7.3). Peptide mapping after digestion with trypsin revealed one major peptide fragment (7.9 kDa) in the DHFR of MAP and MAP/47 and three major tryptic fragments (7.9, 9.6 and 12.5 kDa) in DHFR from MAP/42. We conclude that trimethoprim resistance in H. influenzae results from overproduction of structurally altered DHFR(s).

Chemical Modification of Human Lysozyme. Acetylation and Nitration of Tyrosine

1971 ◽  
Vol 49 (7) ◽  
pp. 816-821 ◽  
Author(s):  
R. L. Fawcett ◽  
T. J. Limbird ◽  
Sandra L. Oliver ◽  
C. L. Borders Jr.
Keyword(s):  
Enzyme Activity ◽  
Chemical Modification ◽  
Gel Filtration ◽  
Degree Of Polymerization ◽  
Tyrosine Residue ◽  
Amino Groups ◽  
Human Lysozyme ◽  
Tyrosine Residues ◽  
Typical Experiment ◽  
High Degree

When human lysozyme is reacted with a 60 M excess of N-acetylimidazole, only one of six tyrosine residues and two amino groups are acetylated. The acetylated lysozyme is 1.2 times as active towards M. lysodeikticus as the unmodified enzyme. When human lysozyme is reacted with a 4 M excess of tetranitromethane, approximately one tyrosine is nitrated. The tetranitromethane also simultaneously induces a high degree of polymerization of the lysozyme. In a typical experiment, nitration leads to a polymerized product that has only 25% of the activity of unmodified enzyme towards M. lysodeikticus. The polymerized lysozyme can be separated into several components by gel filtration on Sephadex G-75. Enzyme activity analyses of the chromatographed lysozyme oligomers indicate that tetranitromethane reduces the activity of human lysozyme primarily by polymerization, since the lysozyme monomer, which contains one nitrotyrosine per molecule, has 65% activity while the trimer has only 5% activity. N-Acetylglucosamine, N,N′-diacetylchitobiose, and N,N',N″-triacetylchitotriose, all inhibitors or substrates of human lysozyme, prevent neither the nitration of the single tyrosine residue nor the polymerization due to tetranitromethane action.

Isolation of Antithrombin III from Normal and α1-Antitrypsin-Deficient Human Plasma

1977 ◽  
Vol 38 (02) ◽  
pp. 0475-0485 ◽  
Author(s):  
Anna D. Borsodi ◽  
Ralph A. Bradshaw
Keyword(s):  
Enzymatic Activity ◽  
Isoelectric Focusing ◽  
Antithrombin Iii ◽  
Factor Xa ◽  
Antithrombin Activity ◽  
Bovine Trypsin ◽  
Normal Plasma ◽  
Antitrypsin Deficiency ◽  
Simplified Procedure ◽  
Stimulation Of

SummaryThe plasma of individuals, hetero- or homozygous for α1-antitrypsin deficiency, contains greatly decreased amounts of antithrombin activity as assayed against factor Xa. However, heparin stimulation of the residual antithrombin activity is observed, which is comparable to that of normal plasma. Antithrombins isolated from both normal and α1-antitrypsin deficient plasma by a simplified procedure are indistinguishable in both properties and yields. The microheterogeneity observed on isoelectric focusing of both preparations can be eliminated by treatment with neuraminidase. Neither purified human antithrombin nor α1-antitrypsin, when assayed against bovine trypsin, is stimulated by heparin. These results clearly establish the unique natures of antithrombin and α1-antitrypsin and show that about 75% of the antithrombin activity measured in normal plasma is due to α1-antitrypsin. Estimates of anti thrombin III activity in normal plasma by assays dependent on enzymatic activity can probably be obtained only in the presence of heparin.

TRIGEMINAL PROJECTIONS IN THE RETICULAR FORMATION OF THE PONS IN THE CAT

1962 ◽  
Vol 40 (1) ◽  
pp. 7-12
Author(s):  
J. M. Langlois ◽  
Guy Lamarche
Keyword(s):  
Reticular Formation ◽  
Trigeminal Nerve ◽  
Unit Activity ◽  
Functional Organization ◽  
Pontine Reticular Formation ◽  
Recording Unit ◽  
Spatial Convergence ◽  
The Face ◽  
High Degree ◽  
Stimulation Of

The projections of the trigeminal nerve in the pontine reticular formation of the cat have been investigated by recording unit activity, after physiological stimulation of the face, in 30 "encéphales isolés" preparations. No somatotopical arrangement was found but a high degree of spatial convergence onto pontine reticular units exists and a certain degree of functional organization was observed.

scholarly journals Purification and subunit structure of argininosuccinate lyase from Chlamydomonas reinhardi

1977 ◽  
Vol 167 (1) ◽  
pp. 71-75 ◽  
Author(s):  
R F Matagne ◽  
J P Schlösser
Keyword(s):  
Enzyme Activity ◽  
Crude Extract ◽  
Enzyme Preparation ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Activity Analysis ◽  
Disc Electrophoresis ◽  
Subunit Structure ◽  
Argininosuccinate Lyase

Argininosuccinate lyase (EC 4.3.2.1) was purified by (NH4)2SO4 fractionation, chromatography on DEAE-cellulose and gel filtration on Sephadex G-200. The final enzyme preparation was purified 46-fold compared with the crude extract. Electrophoresis of this preparation revealed three bands, the major one having the enzyme activity. Analysis of the enzyme by gel filtration and by disc electrophoresis (in two different concentrations of acrylamide) gave mol.wts. of 200000 (+/- 15000) and 190000 (+/- 20000) respectively. Treatment with sodium dodecyl sulphate and mercaptoethanol dissociated the enzyme into subunits of mol.wt. 39000 (+/-2000). The results are indicative of the multimeric structure of the enzyme, which is composed of five (perhaps four or six) identical subunits.

scholarly journals Stimulation of 5-lipoxygenase activity under conditions which promote lipid peroxidation

1989 ◽  
Vol 263 (2) ◽  
pp. 565-572 ◽  
Author(s):  
D Riendeau ◽  
D Denis ◽  
L Y Choo ◽  
D J Nathaniel
Keyword(s):  
Lipid Peroxidation ◽  
High Speed ◽  
Chemical Reduction ◽  
Gel Filtration ◽  
Maximal Activity ◽  
Lipoxygenase Activity ◽  
Supernatant Fraction ◽  
Acid Oxidation ◽  
Thiol Compounds ◽  
Stimulation Of

The characteristics of hydroperoxide activation of 5-lipoxygenase were examined in the high speed supernatant fraction prepared from rat polymorphonuclear leukocytes. Stimulation of 5-lipoxygenase activity by the 5-hydroperoxyeicosatetraenoic acid (5-HPETE) reaction product was strongly dependent on the presence of thiol compounds. Various reducing agents such as mercaptoethanol and glutathione (0.5-2 mM) inhibited the reaction and increased the concentrations of 5-HPETE (1-10 microM) necessary to achieve maximal arachidonic acid oxidation. The requirement for 5-HPETE was not specific and could be replaced by H2O2 (10 microM) but not by the 5-hydroxyeicosatetraenoic acid (5-HETE) analogue. Furthermore, gel filtration chromatography of the soluble extract from leukocytes resolved different fractions which can increase the hydroperoxide dependence or fully replace the stimulation by 5-HPETE. Maximal activity of the 5-HPETE-stimulated reaction required Ca2+ ions (0.2-1 mM) and ATP with the elimination of the HPETE requirement at high ATP concentrations (2-4 mM). In addition, NADPH (1-2 mM), FAD (1 mM), Fe2+ ions (20-100 microM) and chelated Fe3+ (0.1 mM-EDTA/0.1 mM-FeCl3) all markedly increased product formation by 5-lipoxygenase whereas NADH (1 mM) was inhibitory and Fe3+ (20-100 microM) alone had no effect on the reaction. The stimulation by Fe2+ ions and NADPH was also observed under various conditions which increase the hydroperoxide dependence such as pretreatment of the enzyme preparation with glutathione peroxidase or chemical reduction with 0.015% NaBH4. These results provide evidence for an hydroperoxide activation of 5-lipoxygenase which is not product-specific and is modulated by thiol levels and several soluble components of the leukocytes. They also indicate that stimulation of 5-lipoxygenase activity can contribute to increase lipid peroxidation in iron and nucleotide-promoted reactions.

scholarly journals Partial purification and characterization of an endo-α-N-acetylgalactosaminidase from the culture medium of Streptomyces sp. OH-11242

1992 ◽  
Vol 288 (2) ◽  
pp. 475-482 ◽  
Author(s):  
I Ishii-Karakasa ◽  
H Iwase ◽  
K Hotta ◽  
Y Tanaka ◽  
S Omura
Keyword(s):  
Enzyme Activity ◽  
Culture Medium ◽  
Gel Filtration ◽  
Partial Purification ◽  
Gastric Mucus ◽  
Purification And Characterization ◽  
Streptomyces Sp ◽  
Optimum Ph ◽  
New Type

For the purification of a new type of endo-alpha-N-acetylgalactosaminidase from the culture medium of Streptomyces sp. OH-11242 (endo-GalNAc-ase-S) [Iwase, Ishii, Ishihara, Tanaka, Omura & Hotta (1988) Biochem. Biophys. Res. Commun. 151, 422-428], a method for assaying enzyme activity was established. Using purified pig gastric mucus glycoprotein (PGM) as the substrate, oligosaccharides liberated from PGM were pyridylaminated, and the reducing terminal sugars of oligosaccharides larger than Gal beta 1-3GalNAc were analysed by h.p.1.c. The crude enzyme of endo-GalNAc-ase-S was prepared as an 80% (w/v) ammonium sulphate precipitate from the concentrated culture medium. The enzyme was partially purified by gel chromatofocusing and subsequent DEAE-Toyopearl chromatography. Endo-enzyme activity eluted around pI 4.8 on a gel chromatofocusing column and eluted with 0.19-0.25 M-NaCl on a DEAE-Toyopearl column. In the enzyme fraction obtained, no exo-glycosidases or proteases could be detected. The molecular mass of the enzyme was estimated as 105 kDa by gel filtration, and the optimum pH was 5.5. Endo-GalNAc-ase-S hydrolysed the O-glycosidic linkage between GalNAc and Ser (Thr) in 3H-labelled and unlabelled asialofetuin, liberating both the disaccharide (Gal beta 1-3GalNAc) and the tetrasaccharide [Gal beta 1-3 (Gal beta 1-4GlcNAc beta 1-6)GalNAc]. When endo-alpha-N-acetylgalactosaminidase from Alcaligenes sp. (endo-GalNac-ase-A) was incubated with 3H-labelled and unlabelled asialofetuin, only the disaccharide (Gal beta 1-3GalNAc) was liberated.

scholarly journals Interactions Between Chill-Hours and Degree-Days Affect Carpogenic Germination in Monilinia vaccinii-corymbosi

2001 ◽  
Vol 91 (1) ◽  
pp. 77-83 ◽  
Author(s):  
H. Scherm ◽  
A. T. Savelle ◽  
P. L. Pusey
Keyword(s):  
Laboratory Experiments ◽  
Field Experiments ◽  
Degree Days ◽  
Carpogenic Germination ◽  
Conditioning Period ◽  
Low Degree ◽  
Heating Degree Days ◽  
Relationship Of ◽  
High Degree ◽  
The Relationship

The relationship of cumulative chill-hours (hours with a mean temperature <7.2°C) and heating degree-days (base 7.2°C) to carpogenic germination of pseudosclerotia of Monilinia vaccinii-corymbosi, which causes mummy berry disease of blueberry, was investigated. In two laboratory experiments, pseudosclerotia collected from rabbiteye blueberry in Georgia were conditioned at 5 to 6°C for 26 to 1,378 h prior to placement in conditions favorable for germination and apothecium development. The number of chill-hours accumulated during the conditioning period affected the subsequent proportion of pseudosclerotia that germinated and produced apothecia, with the greatest incidence of carpogenic germination occurring after intermediate levels of chilling (≈700 chill-hours). The minimum chilling requirement for germination and apothecium production was considerably lower than that reported previously for pseudo-sclerotia from highbush blueberry in northern production regions. The rate of carpogenic germination was strongly affected by interactions between the accumulation of chill-hours and degree-days during the conditioning and germination periods; pseudosclerotia exposed to prolonged chilling periods, once transferred to suitable conditions, germinated and produced apothecia more rapidly (after fewer degree-days had accumulated) than those exposed to shorter chilling periods. Thus, pseudosclerotia of M. vaccinii-corymbosi are adapted to germinate carpogenically following cold winters (high chill-hours, low degree-days) as well as warm winters (low chill-hours, high degree-days). Results were validated in a combined field-laboratory experiment in which pseudosclerotia that had received various levels of natural chilling were allowed to germinate in controlled conditions in the laboratory, and in two field experiments in which pseudosclerotia were exposed to natural chilling and germination conditions. A simple model describing the timing of apothecium emergence in relation to cumulative chill-hours and degree-days was developed based on the experiments. The model should be useful for better timing of field scouting programs for apothecia to aid in management of primary infection by M. vaccinii-corymbosi.

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