Patterns of development of glycosidase activities in the pea epicotyl

1970 ◽  
Vol 48 (6) ◽  
pp. 1165-1169 ◽  
Author(s):  
Anne Harmon Datko ◽  
G. A. Maclachlan
Keyword(s):  
Indoleacetic Acid ◽  
Specific Activity ◽  
Fresh Weight ◽  
Glucosidase Activity ◽  
Time Treatment ◽  
Glycosidase Activity ◽  
Specific Localization ◽  
Weight Protein ◽  
Pea Epicotyl ◽  
Hydrolysis Of

Rates of hydrolysis of α- and β-linked glucosides and galactosides were compared in reaction mixtures containing nitrophenol derivatives, maltose, or cellobiose as substrates, plus enzyme derived from various parts of the third internode of the pea epicotyl. β-Glucosidase activity (EC. 3.2.1.21) per unit fresh weight, protein, or DNA was concentrated in meristematic tissues (plumule and hook) and nearly absent from adjacent growing and maturing regions in the internode. The other glycosidase activities showed no such specific localization. β-Glucosidase was also the only one of these enzymes that increased in specific activity in the decapitated epicotyl after treatment with indoleacetic acid. The increase (about twofold in 2 days) occurred if cell division was evoked at the same time. Treatment with gibberellic acid had little effect on any glycosidase activity. It is concluded that β-glucosidase activity may have been especially useful in cells during or shortly after cytokinesis, but none of the glycosidase activities were needed specifically for cell elongation or expansion.

STUDIES ON THE REGULATION OF CELLULASE ACTIVITY AND GROWTH IN EXCISED PEA EPICOTYL SECTIONS

1967 ◽  
Vol 45 (10) ◽  
pp. 1837-1844 ◽  
Author(s):  
Der-Fong Fan ◽  
G. A. Maclachlan
Keyword(s):  
Turnover Rate ◽  
Actinomycin D ◽  
Indoleacetic Acid ◽  
Specific Activity ◽  
Cellulase Activity ◽  
Pisum Sativum L ◽  
Pea Epicotyl ◽  
D 20

Applied indoleacetic acid (10−4 – 10−6 M) increased elongation and the amount and specific activity of cellulase in sections detached from the epicotyl of Pisum sativum L. var. Alaska. Actinomycin D (20 μg/ml) inhibited growth and the capacity of sections for synthesizing protein from absorbed 14C-leucine. At the same time it caused cellulase levels to decrease at a rate which indicated a half-life for the enzyme of less than 24 h. Pea cellulase was most stable in vitro between pH 6 and 7; at or below pH 5 its rate of denaturation was comparable to its turnover rate in vivo. Fractionation of sections yielded wall preparations which contained cellulase at a higher specific activity than particles or supernatant. It is concluded that cellulase is synthesized in excised sections by an auxin-regulated mechanism. It is proposed that the enzyme is transported to the wall where it may promote elongation and eventually become denatured.

scholarly journals The Incorporation of Leucine-C14 into Microsomal Particles and other Subcellular Components of the Pea Epicotyl

1959 ◽  
Vol 5 (1) ◽  
pp. 59-68 ◽  
Author(s):  
Paul O. P. Ts'o ◽  
Clifford S. Sato
Keyword(s):  
Specific Activity ◽  
Gradient Centrifugation ◽  
High Specific Activity ◽  
Insoluble Fraction ◽  
Supernatant Fraction ◽  
Group Method ◽  
Pea Seedlings ◽  
Pea Epicotyl ◽  
Hydrolysis Of ◽  
End Group

Incorporation of leucine-C14 into subcellular fractions of the apical section of pea seedlings has been studied as a function of the length of incubation. The specific activity of the microsomes was higher than that of the supernatant for short but not for long incubations, in agreement with observations on other systems. In this developing tissue the nuclei and especially the mitochondria appear to incorporate amino acid very rapidly. An insoluble fraction of the microsome pellet, which is presumably a liponucleoprotein complex, was found to possess, after 1 hour of incubation, a specific activity much greater than that of the purified microsomal particles or the supernatant fraction. Ninety-eight per cent of the leucine-C14 in the purified microsomal particles has been shown to possess bound amino groups, presumably in peptide linkages, by the DNP-end group method. These particles liberate but little peptide or protein of very high specific activity when they are destroyed by removal of Mg or by hydrolysis of RNA. Microsomal particles were fractionated into an RNA fraction and five protein fractions by means of density gradient centrifugation. By this method 95 per cent of the RNA can be separated from 90 per cent of the protein of the particle. Furthermore, the RNA fraction has been shown to contain very little protein of high specific activity. A particular protein fraction which contains the remaining 5 per cent of the RNA, possessed after 1 hour of incubation a specific activity 2 to 9 times higher than the protein of the other fractions.

On the Preparation of Bovine α-Thrombin

1978 ◽  
Vol 39 (01) ◽  
pp. 193-200 ◽  
Author(s):  
Erwin F Workman ◽  
Roger L Lundblad
Keyword(s):  
Immobilized Enzyme ◽  
Catalytic Properties ◽  
Specific Activity ◽  
Guanidine Hydrochloride ◽  
Low Molecular Weight ◽  
Reversible Process ◽  
Gel Beads ◽  
Tissue Thromboplastin ◽  
C 50 ◽  
Hydrolysis Of

SummaryAn improved method for the preparation of bovine α-thrombin is described. The procedure involves the activation of partially purified prothrombin with tissue thromboplastin followed by chromatography on Sulfopropyl-Sephadex C-50. The purified enzyme is homogeneous on polyacrylamide discontinuous gel electrophoresis and has a specific activity toward fibrinogen of 2,200–2,700 N.I.H. U/mg. Its stability on storage in liquid media is dependent on both ionic strenght and temperature. Increasing ionic strength and decreasing temperature result in optimal stability. The denaturation of α-thrombin by guanidine hydrochloride was found to be a partially reversible process with the renatured species possessing properties similar to “aged” thrombin. In addition, the catalytic properties of a-thrombin covalently attached to agarose gel beads were also examined. The activity of the immobilized enzyme toward fibrinogen was affected to a much greater extent than was the hydrolysis of low molecular weight, synthetic substrates.

scholarly journals β-Glucosidase Activity of Lactiplantibacillus plantarum UNQLp 11 in Different Malolactic Fermentations Conditions: Effect of pH and Ethanol Content

Fermentation ◽  
2021 ◽  
Vol 7 (1) ◽  
pp. 22
Author(s):  
Natalia S. Brizuela ◽  
Marina Arnez-Arancibia ◽  
Liliana Semorile ◽  
María Ángeles Pozo-Bayón ◽  
Bárbara M. Bravo-Ferrada ◽  
...  
Keyword(s):  
Pinot Noir ◽  
Gas Chromatography Mass Spectrometry ◽  
Red Wines ◽  
Ph Values ◽  
Glucosidase Activity ◽  
Sorptive Extraction ◽  
Ethanol Content ◽  
Volatile Precursors ◽  
Hydrolysis Of ◽  
High Ethanol

Lactiplantibacillus plantarum strain UNQLp 11 is a lactic acid bacterium with the potential to carry out malolactic fermentation (MLF) in red wines. Recently, the complete genome of UNQLp 11 was sequenced and this strain possesses four loci of the enzyme β-glucosidase. In order to demonstrate that these glucosidase enzymes could be functional under harsh wine conditions, we evaluated the hydrolysis of p-nitrophenyl-β-D-glucopyranoside (p-NPG) in synthetic wine with different ethanol contents (0%, 12%, and 14% v/v) and at different pH values (3.2, 3.5, and 3.8). Then, the hydrolysis of precursor n-octyl β-D-glucopyranoside was analyzed in sterile Pinot Noir wine (containing 14.5% v/v of ethanol, at different pH values) by headspace sorptive extraction gas chromatography-mass spectrometry (HSSE-GC/MS). The hydrolysis of p-NPG showed that β-glucosidase activity is very susceptible to low pH but induced in the presence of high ethanol content. Furthermore, UNQLp 11 was able to release the glycosilated precursor n-octyl, during MLF to a greater extent than a commercial enzyme. In conclusion, UNQLp 11 could improve the aromatic profile of the wine by the release of volatile precursors during MLF.

Characterization of a factor that stimulates hydrolysis of GTP bound to ras gene product p21 (GTPase-activating protein) and correlation of its activity to cell density

1988 ◽  
Vol 8 (10) ◽  
pp. 4169-4173
Author(s):  
M Hoshino ◽  
M Kawakita ◽  
S Hattori
Keyword(s):  
Gene Product ◽  
Cell Density ◽  
Cultured Cells ◽  
Specific Activity ◽  
Gtpase Activating Protein ◽  
Ras Gene ◽  
Cell Extracts ◽  
Mouse Tissues ◽  
Almost All ◽  
Hydrolysis Of

The postmicrosomal fraction of the extract from NIH 3T3 and BALB/c 3T3 cells stimulated the hydrolysis of GTP bound to H-ras gene product p21 by severalfold. The stimulation was observed with normal p21 but not with p21 with valine as the 12th residue. This specificity is similar to that of GTPase-activating protein (GAP) for N-ras p21 described by M. Trahey and F. McCormick (Science 238:542-545, 1987). Consistent with this specificity, analysis of p21-bound nucleotides in living cells revealed that almost all normal p21 bound GDP, whereas oncogenic mutant p21s bound both GTP and GDP. Similar activity was also found in various mouse tissues, with brain tissue showing the highest specific activity. When cell extracts were prepared from cultured cells, there was a linear relationship between GAP activity and cell density. These results suggest the factor is involved in the regulation of cell proliferation.

scholarly journals A Monoacylglycerol Lipase from Mycobacterium smegmatis Involved in Bacterial Cell Interaction

2010 ◽  
Vol 192 (18) ◽  
pp. 4776-4785 ◽  
Author(s):  
Rabeb Dhouib ◽  
Françoise Laval ◽  
Frédéric Carrière ◽  
Mamadou Daffé ◽  
Stéphane Canaan
Keyword(s):  
Mycobacterium Smegmatis ◽  
Wild Type Strain ◽  
Specific Activity ◽  
Cell Interaction ◽  
Monoacylglycerol Lipase ◽  
Homologous Proteins ◽  
Dependent Activity ◽  
Hydrolysis Of

ABSTRACT MSMEG_0220 from Mycobacterium smegmatis, the ortholog of the Rv0183 gene from M. tuberculosis, recently identified and characterized as encoding a monoacylglycerol lipase, was cloned and expressed in Escherichia coli. The recombinant protein (rMSMEG_0220), which exhibits 68% amino acid sequence identity with Rv0183, showed the same substrate specificity and similar patterns of pH-dependent activity and stability as the M. tuberculosis enzyme. rMSMEG_0220 was found to hydrolyze long-chain monoacylglycerol with a specific activity of 143 ± 6 U mg−1. Like Rv0183 in M. tuberculosis, MSMEG_0220 was found to be located in the cell wall. To assess the in vivo role of the homologous proteins, an MSMEG_0220 disrupted mutant of M. smegmatis (MsΔ0220) was produced. An intriguing change in the colony morphology and in the cell interaction, which were partly restored in the complemented mutant containing either an active (ComMsΔ0220) or an inactive (ComMsΔ0220S111A) enzyme, was observed. Growth studies performed in media supplemented with monoolein showed that the ability of both MsΔ0220 and ComMsΔ0220S111A to grow in the presence of this lipid was impaired. Moreover, studies of the antimicrobial susceptibility of the MsΔ0220 strain showed that this mutant is more sensitive to rifampin and more resistant to isoniazid than the wild-type strain, pointing to a critical structural role of this enzyme in mycobacterial physiology, in addition to its function in the hydrolysis of exogenous lipids.

scholarly journals Structural and Catalytic Characterization of TsBGL, a β-Glucosidase From Thermofilum sp. ex4484_79

2021 ◽  
Vol 12 ◽  
Author(s):  
Anke Chen ◽  
Dan Wang ◽  
Rui Ji ◽  
Jixi Li ◽  
Shaohua Gu ◽  
...  
Keyword(s):  
Catalytic Domain ◽  
Specific Activity ◽  
Maximum Activity ◽  
Homologous Proteins ◽  
Glycosidic Bonds ◽  
Catalytic Sites ◽  
Potential Applications ◽  
Beta Glucosidase ◽  
Hydrolysis Of

Beta-glucosidase is an enzyme that catalyzes the hydrolysis of the glycosidic bonds of cellobiose, resulting in the production of glucose, which is an important step for the effective utilization of cellulose. In the present study, a thermostable β-glucosidase was isolated and purified from the Thermoprotei Thermofilum sp. ex4484_79 and subjected to enzymatic and structural characterization. The purified β-glucosidase (TsBGL) exhibited maximum activity at 90°C and pH 5.0 and displayed maximum specific activity of 139.2μmol/min/mgzne against p-nitrophenyl β-D-glucopyranoside (pNPGlc) and 24.3μmol/min/mgzen against cellobiose. Furthermore, TsBGL exhibited a relatively high thermostability, retaining 84 and 47% of its activity after incubation at 85°C for 1.5h and 90°C for 1.5h, respectively. The crystal structure of TsBGL was resolved at a resolution of 2.14Å, which revealed a classical (α/β)8-barrel catalytic domain. A structural comparison of TsBGL with other homologous proteins revealed that its catalytic sites included Glu210 and Glu414. We provide the molecular structure of TsBGL and the possibility of improving its characteristics for potential applications in industries.

Anterior midgut proteinase inhibitor from Glossina morsitans morsitans Westwood (Diptera: Glossinidae) and its effects upon tsetse digestive enzymes

1980 ◽  
Vol 58 (1) ◽  
pp. 79-87 ◽  
Author(s):  
Jon G. Houseman
Keyword(s):  
Proteinase Inhibitor ◽  
Periplaneta Americana ◽  
Specific Activity ◽  
Glossina Morsitans Morsitans ◽  
Tsetse Flies ◽  
Trypsin Activity ◽  
Glossina Morsitans ◽  
Trypsin Hydrolysis ◽  
Hydrolysis Of ◽  
Anterior Midgut

The anterior midgut of Glossina morsitans morsitans Westwood contains a proteinase inhibitor, molecular weight 5000 ± 2000daltons, stable to 1 M HCl, heat, and dialysis, but unstable to 1% trichloroacetic acid. Inhibitor activity is not associated with anticoagulant in the anterior midgut. The specific activity of the proteinase inhibitor is similar in mated and unmated females and greater than in male tsetse flies. Proteinase inhibitor inhibits proteinase VI and trypsin hydrolysis of N-benzoyl-L-arginine ethyl ester (BAEE) and benzoyl-DL-arginine-p-nitroanilide (BAPNA) but has no effect on proteinase VI hydrolysis of haemoglobin. Inhibition of trypsin hydrolysis of haemoglobin is noncompetitive. Proteinase inhibitor levels in the anterior midgut decreased immediately after feeding and then increased, reaching a maximum 60–100 h after ingestion of the bloodmeal. Postteneral flies contained higher levels of proteinase inhibitor than teneral individuals. Trypsin activity in gut homogenates of Phormia regina and Aedes aegypti was inhibited by the tsetse inhibitor. There was no detectable inhibition of bovine or Pterostichus adstrictus trypsin activity. Inhibition of Periplaneta americana trypsin occurred but was less than fly trypsin inhibition. The possible role of the inhibitor in terminating proteinase production is discussed.

scholarly journals Purification and characterization of a heat-stable esterase from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius

1988 ◽  
Vol 250 (2) ◽  
pp. 453-458 ◽  
Author(s):  
H Sobek ◽  
H Görisch
Keyword(s):  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sulfolobus Acidocaldarius ◽  
Sedimentation Equilibrium ◽  
Prolonged Storage ◽  
Heat Stable ◽  
Equilibrium Data ◽  
Poly Ethylene ◽  
Hydrolysis Of

A heat-stable esterase has been purified 1080-fold to electrophoretic homogeneity from Sulfolobus acidocaldarius, a thermoacidophilic archaebacterium; 20% of the starting activity is recovered. The purified enzyme shows a specific activity of 158 units/mg, based on the hydrolysis of p-nitrophenyl acetate. The esterase hydrolyses short-chain p-nitrophenyl esters, aliphatic esters and triacylglycerols. It is strongly inhibited by paraoxon and phenylmethanesulphonyl fluoride, but only weakly by eserine. From sedimentation-equilibrium data and molecular sieving in polyacrylamide gels, the Mr of the esterase is estimated to be 117000-128000. SDS/polyacrylamide-gel electrophoresis reveals a single band of protein, of Mr 32000. The purified esterase crystallizes in the presence of poly(ethylene glycol) in short rods. The enzyme is inactivated only on prolonged storage at temperature above 90 degrees C.

New insights into lactase and glycosylceramidase activities of rat lactase-phlorizin hydrolase

1989 ◽  
Vol 257 (4) ◽  
pp. G616-G623 ◽  
Author(s):  
H. A. Buller ◽  
A. G. Van Wassenaer ◽  
S. Raghavan ◽  
R. K. Montgomery ◽  
M. A. Sybicki ◽  
...  
Keyword(s):  
Active Sites ◽  
Specific Activity ◽  
Fold Increase ◽  
Developmental Pattern ◽  
Lactose Hydrolysis ◽  
Adult Males ◽  
Lactase Activity ◽  
Developmental Patterns ◽  
Catalytic Sites ◽  
Hydrolysis Of

Lactase-phlorizin hydrolase, a small intestinal disaccharidase, has been considered mainly an enzyme important only for the hydrolysis of lactose. After weaning in most mammals lactase-specific activity falls markedly, and, functionally, adult mammals are considered to be lactase deficient. However, the persistence of low levels of lactase activity in adulthood has never been explained. In addition, it has been suggested that lactase-phlorizin hydrolase is associated with glycosylceramidase activity when the enzyme is prepared by column chromatography, but it is unclear whether this represents copurified activities or two catalytic sites on one peptide. The developmental patterns of lactase-phlorizin hydrolase and other disaccharidases were investigated in homogenates of total rat small intestine; lactase and several glycosylceramidases were measured in immunoprecipitates from these homogenates using a monoclonal antibody. The developmental pattern of total lactase activity showed a steady 2.3-fold increase to adult levels (specific activity decreased eightfold), whereas total phlorizin-hydrolase activity increased 10.7-fold (specific activity decreased threefold). As expected, levels of both total and specific sucrase and maltase activities increased during development. In lactating rats total lactase activity showed a significant increase compared with adult males. The developmental pattern of the enzyme activities for the glycolipid substrates was similar to that found for lactase, and the immunoprecipitated enzyme showed a 40- to 55-fold higher affinity for the glycolipids than for lactose. Galactosyl- and lactosylceramide inhibited lactose hydrolysis by 38%, without a competitive pattern, suggesting two different active sites for lactose and glycolipid hydrolysis, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

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