Cloning, expression, characterization and application of protease produced by Bacillus cereus PMW8

Esakkiraj, Palanichamy; Meleppat, Balraj; Lakra, Avinash Kant; Ayyanna, Repally; Arul, Venkatesan

doi:10.1039/c5ra27671c

Cited by 16 publications

(8 citation statements)

References 26 publications

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“…However, complete loss in activity of fibrinolytic protease from Bacillus cereus RSA1 was observed with PMSF (0.99%) and DFP (0.39%), and there was ≈2-fold increase in the activity with β-mercaptoethanol (191.41%) and DTT (183.42%), which indicates it to be a thiol-dependent serine fibrinolytic protease ( Figure 8A). The present study is supported by the protease produced by Bacillus cereus PMW8 [70]. Thiol-dependent protease was also obtained from Exiguobacterium sp.…”

Section: Effect Of Inhibitors/activators Metal Ions and Organic Solsupporting

confidence: 59%

Thrombolytic Potential of Novel Thiol-Dependent Fibrinolytic Protease from Bacillus cereus RSA1

et al. 2019

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The present study demonstrates the production and thrombolytic potential of a novel thermostable thiol-dependent fibrinolytic protease by Bacillus cereus RSA1. Statistical optimization of different parameters was accomplished with Plackett-Burman design and validated further by central composite design with 30.75 U/mL protease production. Precipitation and chromatographic approaches resulted in 33.11% recovery with 2.32-fold purification. The molecular weight of fibrinolytic protease was 40 KDa and it exhibited a broad temperature and pH stability range of 20-80 • C and pH 5-10 with utmost activity at 50 • C and pH 8, respectively. The protease retained its fibrinolytic activity in organic solvents and enhanced the activity in solutions with divalent cations (Mn 2+ , Zn 2+ , and Cu 2+ ). The enzyme kinetics revealed K m and V max values of 1.093 mg/mL and 52.39 µg/mL/min, respectively, indicating higher affinity of fibrinolytic activity towards fibrin. Also, complete inhibition of fibrinolytic activity with DFP and a 2-fold increase with DTT and β-mercaptoethanol indicates its thiol-dependent serine protease nature. MALDI-TOF analysis showed 56% amino acid sequence homology with Subtilisin NAT OS = Bacillus subtilis subsp. natto. The fibrinolysis activity was compared with a commercial thrombolytic agent for its therapeutic applicability, and fibrinolytic protease was found highly significant with absolute blood clot dissolution within 4 h in in vitro conditions. The isolated fibrinolytic protease of Bacillus cereus RSA1 is novel and different from other known fibrinolytic proteases with high stability and efficacy, which might have wide medicinal and industrial application as a thrombolytic agent and in blood stain removal, respectively.Biomolecules 2020, 10, 3 2 of 23 (active protease), converting soluble fibrinogen (glycoprotein) into fibrin polymer (insoluble blood clot) [5]. Fibrin clots are hydrolyzed by plasmin [12], which is stimulated from plasminogen by plasminogen activators (PAs) [13,14]. This natural dynamic equilibrium is disturbed when the process of natural fibrin clot hydrolysis undergoes pathophysiological shambles, leading to formation of fibrin clots. Such clots may cause hindrance in the circulation of blood, causing blockage in blood vessels, ultimately leading to cardiac disorders which are life-threatening [15,16].Approaches such as use of anti-coagulant agents, anti-platelet drugs, fibrinolytic enzymes, and surgical operations are employed for the treatment of thrombosis and to dissolve the blood clots [17]. Further, there are numerous side effects which may occur following the administration of the available anti-thrombotic strategies, as well as the high expense, which limit their scope. The effects of reperfusion, urticaria (allergic reaction), and hemorrhage are the major inimical consequences of such thrombolytic approaches on human health [18,19]. Other after effects include headache, dizziness, ulcers, increased clotting time, nausea and vomiting, etc. Management of thrombosis usin...

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Section: Effect Of Inhibitors/activators Metal Ions and Organic Solsupporting

confidence: 59%

Thrombolytic Potential of Novel Thiol-Dependent Fibrinolytic Protease from Bacillus cereus RSA1

et al. 2019

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“…However, enzyme activity was reduced to 90% by an addition of 10% amstatin. Activity of the protease is not affected by the addition of Tween 20 and Tween 60 (Esakkiraj et al, 2016), while there was an increase in the protease activity after addition of non-ionic detergent Tween 80 (0.2-1% w/v). Padmapriya et al (2012) have also reported an increase in the activity of the protease from marine Bacillus species.…”

Section: Effect Of Surfactants Oxidizing Agents and Detergentsmentioning

confidence: 89%

Characterization of Geobacillus stearothermophilus protease for detergent industry

Iqbal¹,

Aftab²,

Afzal³

et al. 2020

Rev.Mex.Ing.Quim.

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“…Thermostability tests at 50 °C revealed that the activity was the highest after 120 min of heating and relative activity lost only 16% of its value after 240 min ( Figure 8 ). It is not uncommon that relative enzymatic activity increases after short-term heating in the case of thermoactive enzymes, as it happens with protease 32-F38 ( Figure 8 ), and examples of other proteases can be cited [ 48 , 49 ].…”

Section: Resultsmentioning

confidence: 99%

“…They can also act as peptide and ester synthetases and can be used for peptide synthesis and the production of a precursor of the artificial sweetener aspartame [ 58 ], and have been used for the hydrolysis of plant cell walls in order to assist in aqueous extraction processes [ 59 ]. Proteases with similar characteristics and molecular weight also showed anti-biofilm activity against pathogenic bacteria such as Listeria monocytogenes , Escherichia coli , and Salmonella typhi [ 49 ] and are potential candidates for the production of bioactive peptides from casein [ 60 ]. Our initial promising results contribute towards the application of protease 32-F38 in industrial and analytical processes.…”

Section: Discussionmentioning

confidence: 99%

Production and Characterization of an Extracellular Acid Protease from Thermophilic Brevibacillus sp. OA30 Isolated from an Algerian Hot Spring

Gomri¹,

Rico-Díaz

Escuder-Rodríguez

et al. 2018

Microorganisms

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Proteases have numerous biotechnological applications and the bioprospection for newly-thermostable proteases from the great biodiversity of thermophilic microorganisms inhabiting hot environments, such as geothermal sources, aims to discover more effective enzymes for processes at higher temperatures. We report in this paper the production and the characterization of a purified acid protease from strain OA30, a moderate thermophilic bacterium isolated from an Algerian hot spring. Phenotypic and genotypic study of strain OA30 was followed by the production of the extracellular protease in a physiologically-optimized medium. Strain OA30 showed multiple extracellular proteolytic enzymes and protease 32-F38 was purified by chromatographic methods and its biochemical characteristics were studied. Strain OA30 was affiliated with Brevibacillus thermoruber species. Protease 32-F38 had an estimated molecular weight of 64.6 kDa and was optimally active at 50 °C. It showed a great thermostability after 240 min and its optimum pH was 6.0. Protease 32-F38 was highly stable in the presence of different detergents and solvents and was inhibited by metalloprotease inhibitors. The results of this work suggest that protease 32-F38 might have interesting biotechnological applications.

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Cloning, expression, characterization and application of protease produced by Bacillus cereus PMW8

Abstract: Protease enzyme of Bacillus cereus PMW8 possessing antibiofilm activity was cloned and expressed in E.coli BL21(DE3) PLysS.

Cited by 16 publications

References 26 publications

Thrombolytic Potential of Novel Thiol-Dependent Fibrinolytic Protease from Bacillus cereus RSA1

Thrombolytic Potential of Novel Thiol-Dependent Fibrinolytic Protease from Bacillus cereus RSA1

Characterization of Geobacillus stearothermophilus protease for detergent industry

Production and Characterization of an Extracellular Acid Protease from Thermophilic Brevibacillus sp. OA30 Isolated from an Algerian Hot Spring

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