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Journal ArticleDOI

Characterization and biotechnological application of protease from thermophilic Thermomonas haemolytica.

01 Jan 2020-Archives of Microbiology (Arch Microbiol)-Vol. 202, Iss: 1, pp 153-159
TL;DR: According to results obtained from this study, this new strain of Thermomonas haemolytica isolated from geothermal Nenehatun hot spring in Turkey is a promising candidate for industrial applications in production of detergent.
Abstract: In this study, it was aimed to determine the ability to produce protease enzyme of Thermomonas haemolytica isolated from geothermal Nenehatun hot spring in Turkey and utilization of this enzyme in the detergent industry to remove protein stains. The protease-producing strains were screened from hot springs, and a potential strain was identified as T. haemolytica according to morphological, physiological and biochemical characteristics and sequence of 16S rRNA gene. Maximum protease activity was observed at 55 °C and pH 9.0 at 72 h of incubation. Activity was very stable between 50 and 65 °C and pH 8.0–10.0, respectively. The enzyme activity was significantly inhibited by PMSF and partly inhibited by EDTA, EGTA, SDS, and urea. Some divalent metal ions such as Ca2+, Mg2+, and Mn2+ increased the enzyme activity, while Zn2+ and Cu2+ decreased. Michaelis–Menten constant (Km) and maximum velocity (Vmax) values were calculated by Lineweaver–Burk plot as 125 EU/ml and 1262 mg/ml, respectively. The biochemical characterization of the protease obtained from T. haemolytica was performed and applied on the blood and grass-stained fabrics with detergent to evaluate the stain removal performance of the enzyme. It was observed that the application of detergent with enzyme was more effective than the detergent without enzyme to clean up the stained fabrics. This is the first report of characterization of the protease of T. haemolytica. According to results obtained from this study, this new strain is a promising candidate for industrial applications in production of detergent.
Citations
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Journal ArticleDOI
TL;DR: In this paper, the enzyme performance was further optimized to reach 22.903U/mL with an overall optimization fold of 46.7, achieved under the optimized fermentation medium composed of (%) molasses; 8, (NH4)2SO4; 0.45, wheat bran; 5, MgSO4·7H2O; 0., NaCl; 0, NaCl, 0.15, 231.05 and 57.206min−1 respectively.
Abstract: The growing industrial applications of alkaline proteases urged the production of highly active stable enzymes. In the current study, the enzyme production was achieved by submerged fermentation using the bacterial strain Bacillus licheniformis ALW1 that was further optimized to reach 22.903U/mL. The overall optimization fold was 46.7, achieved under the optimized fermentation medium composed of (%) molasses; 8, (NH4)2SO4; 0.45, wheat bran; 5, MgSO4·7H2O; 0.2, KH2PO4; 0.4, NaCl; 0.2 adjusted at pH 8 and incubated for 7days at 37 °C and 280 rpm. Additionally, the enzyme activity after partial purification was optimized by studying the effect of pH and temperature as well as the substrate concentration. The results indicated that the enzyme optimum activity was achieved at pH 9 and 70 °C with Km, Vmax and Kcat values of 3.846 mg/mL, 76.923U/mL/min and 1.206min−1 respectively. Thermal stability study of the partial pure enzyme indicated the half live times of the enzyme as 693.15, 231.05 and 57.76 min−1 at 55, 60 and 65 °C respectively, confirming its thermo-stability. Finally, the efficacy of the partial pure enzyme as a detergent additive was examined. The enzyme retained more than 80% of its activity with an efficient washing performance in the removal of blood stain after 30 min at 50 °C in addition to a commercial detergent.

24 citations

Journal ArticleDOI
TL;DR: In this paper, traditional chemical methods used for shrink resist finishing are suffering from some drawback, which has negative impacts on the performance attributes of the yarn and its performance is not improved by these methods.
Abstract: Felting shrinkage of wool is a major problem which has negative impacts on its performance attributes. Traditional chemical methods used for shrink-resist finishing are suffering from some drawback...

16 citations

Journal ArticleDOI
TL;DR: The results of the study indicated that these bacteria and their enzymes can be used as a source of industrial enzymes.
Abstract: The aim of present study was the isolation and characterization of thermophilic bacteria from three different hot springs in Erzurum, Turkey. For this purpose, 85 bacteria were isolated and characterized by ERIC-PCR genomic fingerprinting and classical identification methods such as morphological, physiological and biochemical characteristics. According to the results, 29 bacterial isolates with different band profiles were analyzed by 16S rRNA gene sequencing and identified as belonging to the genus of Bacillus, Pseudomonas, Silanimonas, Thermomonas and Thauera. This is the first report on the isolation of Silanimonas lenta, Thauera sp. and Thermomonas haemolytica from Turkey Hot Springs. The amylase, lipase and protease enzyme production potentials of the isolates were 80%, 91.25% and 81.25%, respectively. Moreover, 56.47% of the isolates (48) were able to produce all of these enzymes. Therefore, the results of the study indicated that these bacteria and their enzymes can be used as a source of industrial enzymes.

15 citations

Journal ArticleDOI
16 Dec 2020-Biology
TL;DR: The results suggest that this serine protease could clean blood-stained fabrics and showed dehairing activity for cow skin with significantly reduced pollution loads and thus appears to be an eco-friendly additive for a variety of industrial applications.
Abstract: An increased need by the green industry for enzymes that can be exploited for eco-friendly industrial applications led us to isolate and identify a unique protease obtained from a proteolytic Bacillus megaterium-TK1 strain from a seawater source. The extracellular thermostable serine protease was processed by multiple chromatography steps. The isolated protease displayed a relative molecular weight (MW) of 33 kDa (confirmed by zymography), optimal enzyme performance at pH 8.0, and maximum enzyme performance at 70 °C with 100% substrate specificity towards casein. The proteolytic action was blocked by phenylmethylsulfonyl fluoride (PMSF), a serine hydrolase inactivator. Protease performance was augmented by several bivalent metal cations. The protease tolerance was studied under stringent conditions with different industrial dispersants and found to be stable with Surf Excel, Tide, or Rin detergents. Moreover, this protease could clean blood-stained fabrics and showed dehairing activity for cow skin with significantly reduced pollution loads. Our results suggest that this serine protease is a promising additive for various eco-friendly usages in both the detergent and leather industries.

14 citations


Cites background or methods or result from "Characterization and biotechnologic..."

  • ...To study the pH stability, the protease was preincubated in the buffers and its activity was determined as described above [12]....

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  • ...Temperature-dependent protease stability was determined by pre-incubating the protease at temperatures of 20–100 ◦C for 1 h, followed by measuring its remaining activity as described above [12]....

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  • ...In the presence of different metal cations, our protease activity either enhanced (Na+, Mn2+, Ca2+, Mg2+, and Co2+) or remained constant (Cu2+ and Zn2+), except for mercury (Hg2+), which agreed with previous reports [9,12,36]....

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  • ..., arginine, lysine, histidine, and tryptophan) in a protease-active substrate binding site, which results in complete loss of protease activity [9,12,36]....

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  • ...Proteases usually do not tolerate high temperature or surfactants, and thermostable proteases are not able to tolerate alkaline pH conditions or surfactants [4,9,12]....

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Journal ArticleDOI
18 Jan 2021
TL;DR: In this article, a 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.
Abstract: 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.

14 citations

References
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Journal ArticleDOI
TL;DR: On the basis of the assumed theory the rate of the observed reaction is directly proportional to the concentration of the enzyme-substrate compound, where (E:l = (ES).
Abstract: On the basis of the assumed theory the rate of the observed reaction is directly proportional to the concentration of the enzyme-substrate compound, (ES), a t all values of the concentration of the substrate, (S). It is proportional to (S) only a t low values of (S). The numerical value of the dissociation constant is given by the substrate concentration a t half-maximum velocity, where (E:l = (ES). The equilibrium in equation 1 may be heterogeneous or homogeneous. Hitchcock'\" has pointed

11,349 citations

Journal ArticleDOI
TL;DR: In this paper, a review of the proteases that can resist extreme alkaline environments produced by a wide range of alkalophilic microorganisms is presented, and various nutritional and environmental parameters affecting the production of alkaline proteases are delineated.

806 citations

Journal ArticleDOI
TL;DR: This study suggests that Thermus strains play an important role in organic-matter degradation during the thermogenic phase of the composting process.
Abstract: High numbers (10(7) to 10(10) cells per g [dry weight]) of heterotrophic, gram-negative, rod-shaped, non-sporeforming, aerobic, thermophilic bacteria related to the genus Thermus were isolated from thermogenic composts at temperatures between 65 and 82 degrees C. These bacteria were present in different types of wastes (garden and kitchen wastes and sewage sludge) and in all the industrial composting systems studied (open-air windows, boxes with automated turning and aeration, and closed bioreactors with aeration). Isolates grew fast on a rich complex medium at temperatures between 40 and 80 degrees C, with optimum growth between 65 and 75 degrees C. Nutritional characteristics, total protein profiles, DNA-DNA hybridization (except strain JT4), and restriction fragment length polymorphism profiles of the DNAs coding for the 16S rRNAs (16S rDNAs) showed that Thermus strains isolated from hot composts were closely related to Thermus thermophilus HB8. These newly isolated T. thermophilus strains have probably adapted to the conditions in the hot-compost ecosystem. Heterotrophic, ovalspore-forming, thermophilic bacilli were also isolated from hot composts, but none of the isolates was able to grow at temperatures above 70 degrees C. This is the first report of hot composts as habitats for a high number of thermophilic bacteria related to the genus Thermus. Our study suggests that Thermus strains play an important role in organic-matter degradation during the thermogenic phase (65 to 80 degrees C) of the composting process.

228 citations

Journal ArticleDOI
TL;DR: The production of thermostable protease and its characterization in Bacillus species, which is a thermotolerant bacterium, is reported and the presence of galactose and peptone in the medium enhanced enzyme production by 0.5% when compared with other carbon and nitrogen sources.

146 citations