Production of pectin lyase from Geobacillus pallidus p26, purification, characterization and fruit juice application
TL;DR: The purified pectin lyase enzyme was used for getting fruits juices and it was found that yields of fruits juices increased when they were compared with control.
Abstract: A bacterial strain was isolated from Pasinler hot spring, Erzurum, Turkey. The purifi ed thermophilic isolate was identifi ed as Geobacillus pallidus P26 and used to produce extracellular pectin lyase (EC 4.2.2.10). Pectin lyase enzyme was purifi ed 34 fold by using DEAE-cellulose anion exchange column chromatography and characterized. Molecular weight of the enzyme was determined as 56 kDa by using Sephadex G-100 gel fi ltration chromatography. Purifi cation of enzyme was verifi ed by SDS-PAGE. The pH- and temperature optima of enzyme were determined (pH 9.0 and 60 oC, respectively). Pectin lyase was mostly stable at 50 oC for 24 hours. Its' activity decreased to 50 % for 24 h at 60 o C. K M and V max were calculated as 24.8 mg/mL and 2+ but not by Mg 2+ . The purifi ed pectin lyase enzyme was used for getting fruits juices. It was found that yields of fruits juices increased when they were compared with control.
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TL;DR: Investigation of the potential of Penicillium notatum for the production of pectin lyase under solid state culture using wheat bran revealed that pect in lyase isolated from P. notatum is thermally stable and alkaline in nature.
Abstract: Abstract The present study was aimed to investigate the potential of Penicillium notatum for the production of pectin lyase under solid state culture using wheat bran as substrate. Different process parameters were optimized using completely randomized design for enhanced production of the pectin lyase. P. notatum showed maximum production (1875 U/gds) of pectin lyase with substrate amount 15 g/250 ml, moisture level 60%, pH 6, incubation period 120 h at 30°C. Pectin lyase activity was further improved with the addition of maltose and ammonium sulphate as carbon and nitrogen additives (1%), respectively. Partial purification of enzyme was carried out by ammonium sulphate precipitation at 80% saturation level. The P. notatum pectin lyase showed maximal activity at 65°C and pH 8. Km and Vmax values were 0.29% and 0.487 µmol/min, respectively. Energy of activation was found to be 5.33 kJ/mol. A detailed kinetic study of thermal inactivation was carried out. The results showed that pectin lyase exhibited resistance against thermal unfolding. Effect of various metals on pectin lyase activity was also investigated. All the metals showed inhibitory effect on the enzyme activity. The present investigation revealed that pectin lyase isolated from P. notatum is thermally stable and alkaline in nature.
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TL;DR: In this article, a thermophilic bacterium isolated from a thermal source in Turkey and identified as Geobacillus pallidus P26 was used to produce the alkaline phosphatase (ALP) enzyme.
Abstract: In this study, a thermophilic bacterium isolated from a thermal source in Turkey and identified as Geobacillus pallidus P26 was used to produce the alkaline phosphatase (ALP) enzyme. The enzyme was partially isolated by salting out the supernatant with saturated ammonium sulphate. Then, the enzyme was purified 23 times using ion-exchange (DEAE-Sepharose) and size-exclusion chromatography (Sephacryl S-200). Using p-nitrophenyl phosphate (pNPP) substrate, the maximum reaction velocity (Vmax) of the purified enzyme was calculated as 0.255 µmol/min/mg and Michaelis-Menten constant (Km) for pNPP (p-nitrophenyl phosphate) as 3.92 mM. kcat (turnover number) and kcat/Km were calculated as 0.812 s–1 and 207.14 s–1 M–1, respectively. The half-life of the enzyme (t1/2) purified from Geobacillus pallidus G26 was calculated as 38 hours at 60°C, 14 hours at 70°C and 1.5 hours at 80°C. The thermodynamic parameters of ALP (
$$\Delta S_{{{\text{IN}}}}^{{{\# }}}{\text{,}}$$
$$\Delta H_{{{\text{IN}}}}^{\# }$$
and $$\Delta G_{{{\text{IN}}}}^{\# }$$
) were also calculated. The enzymes extracted from thermophilic bacteria can maintain their stability at high temperatures for a longer period. This characteristic is favorable for implementations that require high temperatures.
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TL;DR: Pectinases are one of the most widely distributed enzymes in bacteria, fungi and plants as discussed by the authors, and they have a share of 25% in the global sales of food enzymes.
Abstract: Pectinases or petinolytic enzymes, hydrolyze pectic substances. They have a share of 25% in the global sales of food enzymes. Pectinases are one of the most widely distributed enzymes in bacteria, fungi and plants. Protopectinases, polygalacturonases, lyases and pectin esterases are among the extensively studied pectinolytic enzymes. Protopectinases catalyze the solubilization of protopectin. Polygalacturonases hydrolyze the polygalacturonic acid chain by addition of water and are the most abundant among all the pectinolytic enzymes. Lyases catalyze the trans-eliminative cleavage of the galacturonic acid polymer. Pectinesterases liberate pectins and methanol by de-esterifying the methyl ester linkages of the pectin backbone. Pectinolytic enzymes are of significant importance in the current biotechnological era with their all-embracing applications in fruit juice extraction and its clarification, scouring of cotton, degumming of plant fibers, waste water treatment, vegetable oil extraction, tea and coffee fermentations, bleaching of paper, in poultry feed additives and in the alcoholic beverages and food industries. The present review mainly contemplates on the types and structure of pectic substances, the classification of pectinolytic enzymes, their assay methods, physicochemical and biological properties and a bird's eye view of their industrial applications.
975 citations
"Production of pectin lyase from Geo..." refers background in this paper
...…those are fruit juice processing, coffee and tea processing, macerating of plants and vegetable tissue, degumming of plant fibers, treatment waste water, extracting vegetable oil, bleaching of paper, adding poultry feed and the textile, alcoholic beverages, and food industries (Jayani et al. 2005)....
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TL;DR: The idea is now emerging that this type of yeast enzyme could offer an alternative to fungal enzymes for industrial applications.
Abstract: When grown in the appropriate medium, several yeast species produce pectinases able to degrade pectic substances. It is mainly exocellular endopolygalacturonases that break pectins or pectate down by hydrolysis of α-1,4-glycosidic linkages in a random way. Biochemical characterisation of these enzymes has shown that they have an optimal pH in the acidic region and an optimal temperature between 40 and 55°C. Their production by yeasts is a constitutive feature and is repressed by the glucose concentration and aeration. Pectic substances and their hydrolysis products are used as carbon sources by a limited number of yeasts and hence these enzymes must be involved in the colonisation of different parts of plants, including fruits. The first yeast pectic enzyme (encoded by the PSE3 gene) was cloned from Tichosporon penicillatum. Recently, a polygalacturonase-encoding gene from Saccharomyces cerevisiae has been cloned and overexpressed in several strains and the gene for an extracellualar endopolygalacturonase from Kluyveromyces marxianus has also been described. Taking all the results together, the idea is now emerging that this type of yeast enzyme could offer an alternative to fungal enzymes for industrial applications.
231 citations
"Production of pectin lyase from Geo..." refers background in this paper
...Pectinases are produced by a large number of organisms, such as bacteria (Kim et al. 1998), fungi (Alana et al. 1991, Obi et al. 1985), actinomycetes (Beg et al. 2000) and yeast (Blancoa et al. 1999)....
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