Purification and Characterization of a Membrane-Unbound Highly Thermostable Metalloprotease from Aeromonas Caviae

Santhalembi, Laishram; Pennathur, Gautam

doi:10.1007/s13369-015-1849-9

Cited by 3 publications

(2 citation statements)

References 25 publications

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“…Cellulase, xylanase, and β-1,3-glucanase were produced in solid-state fermentation (SSF) by a strain of Tricoderma citrinoviride AUKAR04 and the enzymes cocktail were partially purified by three-phase partitioning (TPP) method, then redissolved in 25 mL of 10 mM sodium acetate buffer (pH 5.0), and finally, dialyzed against 2 mM sodium acetate buffer (pH 5.0) for 10 h at 4 °C. The obtained enzyme solution was stored in a refrigerator. , The activities of cellulase, xylanase, and β-1,3-glucanase were assayed and the amounts of reducing sugar liberated was estimated by the method adopted in our previous studies. , In this connection, producing enzymes at laboratory scale is a major advantage and minimizes the procurement costs of the enzymes. The total protein concentration was measured by the Bradford’s method with bovine serum albumin as the standard …”

Section: Methodsmentioning

confidence: 99%

“…The obtained enzyme solution was stored in a refrigerator. 19,20 The activities of cellulase, xylanase, and β-1,3-glucanase were assayed and the amounts of reducing sugar liberated was estimated by the method adopted in our previous studies. 21,22 In this connection, producing enzymes at laboratory scale is a major advantage and minimizes the procurement costs of the enzymes.…”

Section: ■ Experimental Sectionmentioning

confidence: 99%

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Bioconversion of Lignocellulosic Biomass to Fermentable Sugars by Immobilized Magnetic Cellulolytic Enzyme Cocktails

et al. 2018

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Enzyme cocktails of reusable, highly stable cellulolytic enzymes play an inevitable role in bioconversion of biomass to biofuels economically. Cellulase, xylanase and β-1,3-glucanase bound silica-amine functionalized iron oxide magnetic nanoparticles (ISN-CLEAs) were prepared and used as the biocatalyst for the depolymerization of cellulosic biomass into monomeric sugar in the present study. The FeO-NPs and FeO@SiO-NH-NPs and ISN-CLEAs had an average hydrodynamic size of 82.2, 86.4, and 976.9 nm, respectively, which was confirmed by dynamic light scattering (DLS). About 97% of protein binding was achieved with 135 mM glutaraldehyde at 10 h of cross-linking time and successful binding was confirmed by Fourier transform infrared spectroscopy (FTIR). The ISN-CLEAs exhibited the highest thermal stability of 95% at 50 °C for 2 h and retained extended storage stability of 97% compared to 60% of its free counterpart. Besides, cross-linking allowed ISN-CLEAs reuse for at least eight consecutive cycles retaining over 70% of its initial activity. ISN-CLEAs exhibited approximately 15% increase in carbohydrate digestibility on sugar cane bagasse and eucalyptus pulp than the free enzyme.

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Section: Methodsmentioning

confidence: 99%

Section: ■ Experimental Sectionmentioning

confidence: 99%

Bioconversion of Lignocellulosic Biomass to Fermentable Sugars by Immobilized Magnetic Cellulolytic Enzyme Cocktails

et al. 2018

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Structural-genetic insight and optimization of protease production from a novel strain of Aeromonas veronii CMF, a gut isolate of Chrysomya megacephala

et al. 2021

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A novel alkali and thermotolerant protease from Aeromonas spp. retrieved from wastewater

Sodagar,

Jalal,

Najafi

et al. 2024

Sci Rep

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Enzymes are integral to numerous industrial processes, with a growing global demand for various enzyme types. Protease enzymes, in particular, have proven to be cost-effective, stable, and compatible alternatives to traditional chemical processes in both industrial and environmental applications. In this study, an alkaline protease-producing strain of Aeromonas spp. was isolated from a wastewater treatment plant in Iran. The protease production was confirmed by culturing the strain on casein agar medium. The bacterium was identified through morphological, biochemical, and 16 S rRNA sequencing analyses. The optimal culture medium for bacterial growth and enzyme production was obtained using peptone, salt, yeast extract, galactose, and CaCl₂ at an initial pH of 8. Maximum protease production was achieved after 20 h of incubation at 40 °C. To partially purify the enzyme, the supernatant of the bacterial culture medium was first centrifuged, and the enzyme was precipitated using ammonium sulfate, followed by dialysis. Zymography revealed the production of one type of protease during bacterial growth. The partially purified protease exhibited optimal activity at pH 8.5 and maximum stability at pH 9. The optimum temperature for maximum enzyme activity was observed at 50 °C, with 100% residual activity retained for 1 h at 0 °C. The effect of metal ions on enzyme activity was assessed, revealing that KCl induced the most significant effects (p < 0.0001) on enzyme activity. Chemical amino acid modifiers and inhibitors, such as EDTA, DEPSI, and IAA, did not exhibit significant inhibition. In contrast, PMSF and HNBB significantly (p < 0.0001) reduced enzyme activity, suggesting that the enzyme could be classified as a serine protease. The protease also demonstrated high stability in the presence of 2% SDS, showing no signs inactivation. The alkaline pH optimum, thermal stability, and resistance to SDS exhibited by the protease produced by the Aeromonas strain are particularly promising characteristics that warrant further investigation. Based on preliminary tests and the enzyme’s characteristics, this protease can be recommended for various applications, pending further studies.

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Purification and Characterization of a Membrane-Unbound Highly Thermostable Metalloprotease from Aeromonas Caviae

Cited by 3 publications

References 25 publications

Bioconversion of Lignocellulosic Biomass to Fermentable Sugars by Immobilized Magnetic Cellulolytic Enzyme Cocktails

Bioconversion of Lignocellulosic Biomass to Fermentable Sugars by Immobilized Magnetic Cellulolytic Enzyme Cocktails

Structural-genetic insight and optimization of protease production from a novel strain of Aeromonas veronii CMF, a gut isolate of Chrysomya megacephala

A novel alkali and thermotolerant protease from Aeromonas spp. retrieved from wastewater

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