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Journal ArticleDOI

Studies on pH and thermal deactivation of pectolytic enzymes from Aspergillus niger

01 Oct 2003-Biochemical Engineering Journal (Elsevier)-Vol. 16, Iss: 1, pp 57-67
TL;DR: The addition of protein doubled the half-life times of partially purified PMG, PG I and PG II and the interaction effect of pectolytic enzymes on deactivation was found to be significant.
About: This article is published in Biochemical Engineering Journal.The article was published on 2003-10-01. It has received 108 citations till now.
Citations
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Journal ArticleDOI
TL;DR: Thermodynamic parameters of ficin produced by Ficus johannis indicate that Ficin from FicusJohannis may have potential applications in the industrial biotechnology.
Abstract: Ficin from Ficus johannis was purified to homogeneity by (NH4)2SO4 precipitation, CM-Sepharose and SP-Sepharose chromatography, and gel filtration Sephadex G-25 chromatography. The enzyme, that is characterized by a molecular weight of 25 kDa measured by SDS-PAGE, showed higher affinity towards casein and the highest catalytic efficiency at pH 7.0 and 60 °C. Catalytic hydrolysis rate follows the Michaelis–Menten equation and values of 32.9 s−1 and 13.7 µM for kcat and Km respectively, by Michaelis–Menten, were found. The pKa1 and pKa2 values of the active site ionizable groups controlling Vmax were 4.2 and 8.7, respectively. Thermodynamic parameters for soluble casein hydrolysis and for irreversible inactivation of ficin at different temperatures in the range from 45 to 60 °C were also determined. This is the first report on the thermodynamic parameters of ficin produced by Ficus johannis. These results indicate that Ficin from Ficus johannis may have potential applications in the industrial biotechnology.

19 citations

Journal ArticleDOI
TL;DR: This enzyme exhibited a t1/2 of 63 min at 60°C and its specific activity, turnover number, and catalytic efficiency were 6.17 U/mg, 113.64 min−1, and 35.18 mL/(min·mg), respectively, revealing a potential application for the industry.
Abstract: For the first time, a polygalacturonase from the culture broth of Tetracoccosporium sp. was isolated and incubated at 30 degrees C in an orbital shaker at 160 rpm for 48 h. The enzyme was purified by ammonium sulfate precipitation and two-step ion-exchange chromatography and had an apparent molecular mass of 36 kDa, as shown by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Its optimum activity was at pH 4.3 and 40 degrees C, and the Km and Vmax values of this enzyme (for polygalacturonic acid) were 3.23 mg/mL and 0.15 micromol/min, respectively. Ag+, Co2+, EDTA, Tween-20, Tween-80, and Triton X-100 stimulated polygalacturonase activity whereas Al3+, Ba2+, Ca2+, Fe2+, Fe3+, Ni2+, Mg2+, Mn2+, and SDS inhibited it. In addition, iodoacetamide and iodoacetic acid did not inhibit enzyme activity at a concentration of 1 mM, indicating that cysteine residues are not part of the catalytic site of polygalacturonase. We studied the kinetic properties and thermal inactivation of polygalacturonase. This enzyme exhibited a t1/2 of 63 min at 60 degrees C and its specific activity, turnover number, and catalytic efficiency were 6.17 U/mg, 113.64 min-1, and 35.18 mL/(min.mg), respectively. The activation energy (DeltaE#) for heat inactivation was 5.341 kJ/mol, and the thermodynamic activation parameters DeltaG#, DeltaH#, and DeltaS# were also calculated, revealing a potential application for the industry.

19 citations

Journal ArticleDOI
TL;DR: In this article, the dry residue of passion fruit (Passiflora edulis) and subsequent characterization of the enzyme polygalacturonase produced by solid state fermentation and leavening agent and the filamentous fungus Aspergillus niger were investigated.
Abstract: Pectinolytic enzymes, or pectinases, are a heterogeneous group of enzymes that hydrolyze the pectic substances present in plant cells. These enzymes are applied in several areas, making it important to know their characteristics for an efficient application. This work aimed to characterize the dry residue of passion fruit (Passiflora edulis) and subsequent characterization of the enzyme polygalacturonase produced by solid state fermentation and leavening agent and the filamentous fungus Aspergillus niger. The residue of passion fruit presented a pectin content of 13.10%, having a potential as a substrate for the production of pectinases. The activity of polygalacturonase reached a maximum value after 66 h of process with 40% initial moisture content and 1% of nitrogen supplementation. In this condition an activity of 20.9 U g-1 was obtained. The polygalacturonase produced the crude enzymatic extract presenting good thermal stability up to temperatures of 50 °C. This enzyme remained stable in the pH range between 3.5 and 5.5 and was not detected for pH values above 6.5.

18 citations

Journal ArticleDOI
TL;DR: Antioxidant capacity and antibacterial activity against Bacillus cereus of enzyme-hydrolyzed milk and permeate from membrane were confirmed and its positive effects, with respect to higher permeate flux and lower energy consumption in filtration process, were proven.
Abstract: Enzymatic hydrolysis of soybean milk proteins with cysteine protease papain was performed in an advanced bioreactor, operated with batch mode. In soybean milk protein hydrolysis reaction, enzyme and substrate ratio and reaction temperature were varied, ranging from 0.029:100-0.457:100 and 30-60 °C, respectively. The degree of hydrolysis of soybean milk proteins was increased with increase of enzyme and substrate (soybean milk protein) ratio. However, the degree of hydrolysis was increased due to change of reaction temperature from 30 °C to 60 °C with enzyme and substrate ratio 0.229:100 and was reduced when hydrolysis reaction was performed with enzyme and substrate ratio 0.11:100 at hydrolysis temperature 60 °C. Antioxidant capacity of enzyme-treated milk had a similar trend with degree of hydrolysis. In a later exercise, a membrane bioreactor was adopted for continuous production of antioxidant and antibacterial peptides from soybean milk. The membrane bioreactor was operated for 12 h with constant feeding. Ceramic-made tubular membrane with a pore size 20 nm was used. Application of static turbulence promoter in a membrane separation process was investigated and its positive effects, with respect to higher permeate flux and lower energy consumption in filtration process, were proven. Antioxidant capacity and antibacterial activity against Bacillus cereus of enzyme-hydrolyzed milk and permeate from membrane were confirmed.

17 citations


Cites background from "Studies on pH and thermal deactivat..."

  • ...Other possible reasons of inactivation of an enzyme are compaction of the enzyme molecule, which may occurr by the generation of charged molecules around the enzyme and dissociation of enzyme structure at high temperatures [8,56]....

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Journal ArticleDOI
TL;DR: The nanoZrO2-CeO2 (150m2/g) and nano ZrO 2-B (296m2 /g) materials were synthesized and characterized on the basis of zirconia as discussed by the authors, and they proved to be less effective supports for a lipase from Candida rugosa.
Abstract: We synthesized and characterized three novel materials on the basis of zirconia. Despite their three-to six fold higher specific surface area, nanoZrO2-CeO2 (150 m2/g) and nanoZrO2-B (296 m2/g) they proved to be less effective supports for a lipase from Candida rugosa than nanoZrO2-A. For the last, we achieved protein loading of 23 mg/g, distributed in a monolayer, which means that 61% of the carrier surface was occupied by the protein. The immobilized on nanoZrO2-A lipase (nanoZrO2-A-CRL) exhibited enhanced thermal and solvent stability in comparison to the native enzyme. NanoZrO2-A-CRL has a half-life at 55 °C of 73 min, while for the native enzyme it was only 5.3 min. The immobilized enzyme preserved 20% of its activity in six consecutive cycles of the reaction hydrolysis of tributyrin. The immobilization influenced the enantioselectivity of CRL. Using nanoZrO2-A-CRL under optimal reaction condition in the reaction of esterification of lauric acid with (±)-menthol, we achieved menthol conversion of 43% (i.e. 82% of (−) methyl laurate), enantiomeric excess of 97% and enantiomeric ratio of 144. The conformational analysis proved that upon immobilization no serious change in the secondary structure of the lipase from Candida rugosa had occurred.

16 citations

References
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Journal Article
TL;DR: Procedures are described for measuring protein in solution or after precipitation with acids or other agents, and for the determination of as little as 0.2 gamma of protein.

289,852 citations

Journal ArticleDOI
TL;DR: In this paper, the probability of the activated state is calculated using ordinary statistical mechanics, and the probability multiplied by the rate of decomposition gives the specific rate of reaction, and necessary conditions for general statistical treatment to reduce to the usual kinetic treatment are given.
Abstract: The calculation of absolute reaction rates is formulated in terms of quantities which are available from the potential surfaces which can be constructed at the present time. The probability of the activated state is calculated using ordinary statistical mechanics. This probability multiplied by the rate of decomposition gives the specific rate of reaction. The occurrence of quantized vibrations in the activated complex, in degrees of freedom which are unquantized in the original molecules, leads to relative reaction rates for isotopes quite different from the rates predicted using simple kinetic theory. The necessary conditions for the general statistical treatment to reduce to the usual kinetic treatment are given.

4,718 citations

Book ChapterDOI

3,073 citations

Journal ArticleDOI
TL;DR: This review discusses various types of pectinases and their applications in the commercial sector.

1,001 citations