Production of alkaline protease by rhizospheric Bacillus cereus HP_RZ17 and Paenibacillus xylanilyticus HP_RZ19

Jadhav, H. P.; Sonawane, Mahesh S.; Khairnar, Mitesh; Sayyed, R. Z.

doi:10.1007/s42398-020-00096-z

Cited by 37 publications

(22 citation statements)

References 31 publications

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“…The concentration of protein (mg/mL) was estimated following Lowry et al (1951) and determined as a measure of its absorbance at 700 nm after each step of the enzyme purification process, using bovine serum albumin (BSA) as a reference [42]. Protease activity of the enzyme (U/mL) was assessed as per standard protocol by Cupp-Enyard, [43] with modifications, as pH and temperature of the assay were optimized wherein the enzyme was incubated at different pH (6)(7)(8)(9)(10)(11)(12) with temperature ranging 10-90 • C. Thus, at each purification process as well as for characterization of enzyme, the protease activity was measured at a standardized pH, 8 and temperature, 70 • C during this study.…”

Section: Protein Estimation and Protease Activity Assaymentioning

confidence: 99%

“…pH effects on the enzymatic activity were assessed using 0.1 M buffer systems comprising of phosphate buffer (pH 6-7), Tris-HCl buffer (pH 8-9) and glycine-NaOH buffer (pH 10-12), whereas the pH stability analysis was conducted by prior incubation of protease enzyme for 1 h at 35 ± 2 • C in pH buffers (6)(7)(8)(9)(10)(11)(12), and residual activities were analyzed.…”

Section: Gel Filtration Column Chromatography and Gel Electrophoresis (Sds-page)mentioning

confidence: 99%

“…Protease enzymes being among the three major industrial enzyme groups, make up for around 65% of the gross enzyme trading globally [ 8 ] and are envisaged in the development of several eco-friendly bio-remedial technologies [ 9 , 10 ]. Although a wide variety of soil microbes produce alkaline proteases [ 11 , 12 ], one of the main drawbacks of these proteases is their unstable nature in alkaline pH at higher temperatures [ 13 ] as these conditions are deleterious for most of the enzymes. Thus, there is a need to prospect new heat-stable alkaline proteases having novel properties owing to growing enzyme demand with better stability [ 14 ].…”

Section: Introductionmentioning

confidence: 99%

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Biochemical Characterization and Functional Analysis of Heat Stable High Potential Protease of Bacillus amyloliquefaciens Strain HM48 from Soils of Dachigam National Park in Kashmir Himalaya

et al. 2021

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A novel temperature stable alkaline protease yielding bacteria was isolated from the soils of Dachigam National Park, which is known to be inhabited by a wide variety of endemic plant and animal species of Western Himalaya. This high-potential protease producing isolate was characterized and identified as Bacillus amyloliquefaciens strain HM48 by morphological, Gram’s staining and biochemical techniques followed by molecular characterization using 16S rRNA approach. The extracellular protease of B. amyloliquefaciens HM48 was purified by precipitating with ammonium sulfate (80%), followed by dialysis and Gel filtration chromatography increasing its purity by 5.8-fold. The SDS–PAGE analysis of the purified enzyme confirmed a molecular weight of about ≈25 kDa. The enzyme displayed exceptional activity in a broad temperature range (10–90 °C) at pH 8.0, retaining its maximum at 70 °C, being the highest reported for this proteolytic Bacillus sp., with KM and Vmax of 11.71 mg/mL and 357.14 µmol/mL/min, respectively. The enzyme exhibited remarkable activity and stability against various metal ions, surfactants, oxidizing agent (H2O2), organic solvents and displayed outstanding compatibility with widely used detergents. This protease showed effective wash performance by exemplifying complete blood and egg-yolk stains removal at 70 °C and efficiently disintegrated chicken feathers making it of vital importance for laundry purpose and waste management. For functional analysis, protease gene amplification of strain HM48 yielded a nucleotide sequence of about 700 bp, which, when checked against the available sequences in NCBI, displayed similarity with subtilisin-like serine protease of B. amyloliquefaciens. The structure of this protease and its highest-priority substrate β-casein was generated through protein modeling. These protein models were validated through futuristic algorithms following which protein–protein (protease from HM48 and β-casein) docking was performed. The interaction profile of these proteins in the docked state with each other was also generated, shedding light on their finer details. Such attributes make this thermally stable protease novel and suitable for high-temperature industrial and environmental applications.

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Section: Protein Estimation and Protease Activity Assaymentioning

confidence: 99%

Section: Gel Filtration Column Chromatography and Gel Electrophoresis (Sds-page)mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Biochemical Characterization and Functional Analysis of Heat Stable High Potential Protease of Bacillus amyloliquefaciens Strain HM48 from Soils of Dachigam National Park in Kashmir Himalaya

et al. 2021

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“…The source and concentration of nitrogen and carbon are crucial aspects to achieve the maximum production of any enzyme as the various physiological pathways require the carbon as a substrates regulates the enzyme production [32]. In present study, the maximum amylase production was observed 1.0% soluble starch as sole source of carbon when compared to other carbon sources tested for isolate Bacillus sp.…”

Section: Discussionmentioning

confidence: 57%

Statistical-Based Bioprocess Optimization of Amylase Production from Halophilic Bacillus sp. H7.

Bandal¹,

Tile²,

Sayyed³

et al. 2021

Preprint

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Using the above results from RMS analysis the optimum values were predicted for the independent significant variables (Figure 3) the optimized levels of these variables in combination with other media variables the maximum production was predicted to be 199.90 U/mL. The predicted data were validated through confirmatory experiments performed in triplicates. A 1.29-fold increase in amylase activity against un-optimized (OVAT) medium was achieved in the present study authenticating the efficacy of RSM in process optimization (Figure 4). 2.6 Model validation and scale-up at laboratory scale (5L) bioreactor Once the parameters were standardized in the shake-flasks culture, the experiment was scaled-up to a laboratory-scale bioreactor (5 L). The yield of amylase increased by 1.01 fold (205.69 U/mL), it could be possible because the enzyme production in a bioreactor is higher than in shake-flasks culture as the various critical variable factors such as the dissolved oxygen (DO) and the pH can be optimally controlled at the desired levels [22].

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“…Paenibacillus was originally included in the genus of Bacillus , and later officially reclassified in 1993 as an individual genus [ 30 ]. Several important beneficial products of this genus have been discovered, including extra-cellular enzymes [ 9 , 10 , 31 ], antimicrobial substances [ 25 , 32 ], exopolysaccharides [ 25 , 33 ], anti-diabetic compounds [ 26 , 27 ], and antioxidants [ 25 , 28 ]. Interestingly, the production of proteases by Paenibacillus strains and their applications are rarely reported.…”

Section: Introductionmentioning

confidence: 99%

Utilization of Seafood Processing By-Products for Production of Proteases by Paenibacillus sp. TKU052 and Their Application in Biopeptides’ Preparation

et al. 2020

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Microbial fermentation of by-products is a renewable and efficient technique in the development of a range of useful products. In this study, protease synthesis by Paenibacillus sp. TKU052 was carried out on culture media containing some common seafood processing by-products (SPBPs) as the sole source of carbon and nitrogen (C/N). The most suitable C/N nutrition source for the production of proteases was found to be 3.0% (w/v) demineralized crab shells powder (deCSP) and maximal enzyme activity of 4.41 ± 0.16 U/mL was detected on the third day of the culture. Two proteases (P1 and P2) with a similar molecular weight of 31 kDa were successfully isolated and purified from the 3-day deCSP-containing medium. Both P1 and P2 exhibited the highest activity of gelatin hydrolysis at pH 6 and 60 °C. The gelatin hydrolysates catalyzed by Paenibacillus TKU052 proteases were evaluated for biological activities, including 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, angiotensin-I converting enzyme (ACE) inhibition, and prebiotic activities. The gelatin hydrolysates expressed 31.76–43.95% DPPH radical scavenging activity and 31.58–36.84% ACE inhibitory activity, which was higher than those from gelatin. Gelatin hydrolysates also showed the growth-enhancing effect on Bifidobacterium bifidum BCRC 14615 with an increase to 135.70–147.81%. In short, Paenibacillus sp. TKU052 could be a potential strain to utilize crab shell wastes to produce proteases for bio-active peptides’ preparation.

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Production of alkaline protease by rhizospheric Bacillus cereus HP_RZ17 and Paenibacillus xylanilyticus HP_RZ19

Cited by 37 publications

References 31 publications

Biochemical Characterization and Functional Analysis of Heat Stable High Potential Protease of Bacillus amyloliquefaciens Strain HM48 from Soils of Dachigam National Park in Kashmir Himalaya

Biochemical Characterization and Functional Analysis of Heat Stable High Potential Protease of Bacillus amyloliquefaciens Strain HM48 from Soils of Dachigam National Park in Kashmir Himalaya

Statistical-Based Bioprocess Optimization of Amylase Production from Halophilic Bacillus sp. H7.

Utilization of Seafood Processing By-Products for Production of Proteases by Paenibacillus sp. TKU052 and Their Application in Biopeptides’ Preparation

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