Medium optimization and immobilization of purified fibrinolytic URAK from Bacillus cereus NK1 on PHB nanoparticles

Deepak, Venkataraman; Ilangovan, Shailaja; Muraleedharan, V. R.; Victoria, Maria; Pasha, Sheik Pran Babu Sardar; Pandian, Sureshbabu Ram Kumar; Gurunathan, Sangiliyandi

doi:10.1016/j.enzmictec.2010.07.004

Cited by 35 publications

(11 citation statements)

References 32 publications

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“…The complete design consisted of 12 experiments with four replicates at the central point carried out in random order. The Response Surface Methodology (RSM) was used to better visualize and compare results [18][19][20][21].…”

Section: Determination Of the Optimal Conditions For Proteases Purifimentioning

confidence: 99%

“…Among these, the Response Surface Methodology (RSM) is a collection of mathematical and statistical techniques that are useful for modeling and analysis in applications where a response of interest (or output) is influenced by several factors. The objective of RSM is to optimize such a response [18][19][20] through experimental design, model fitting, validation and optimization steps, using a minimum number of experiments for a large number of factors. It was already applied successfully to improve the production and/or purification of several enzymes [21,22].…”

Section: Introductionmentioning

confidence: 99%

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Purification of a fibrinolytic protease from Mucor subtilissimus UCP 1262 by aqueous two-phase systems (PEG/sulfate)

Nascimento

Sales

Porto

et al. 2016

Journal of Chromatography B

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A fibrinolytic protease from M. subtilissimus UCP 1262 was recovered and partially purified by polyethylene glycol (PEG)/sodium sulfate aqueous two-phase systems (ATPS). The simultaneous influence of PEG molar mass, PEG concentration and sulfate concentration on the enzyme recovery was first investigated using a 2(3) full factorial design, and the Response Surface Methodology used to identify the optimum conditions for enzyme extraction by ATPS. Once the best PEG molar mass for the process had been selected (6000g/mol), a two-factor central composite rotary design was applied to better evaluate the effects of the other two independent variables. The fibrinolytic enzyme was shown to preferentially partition to the bottom phase with a partition coefficient (K) ranging from 0.2 to 0.7. The best results in terms of enzyme purification were obtained with the system formed by 30.0% (w/w) PEG 6000g/mol and 13.2% (w/w) sodium sulfate, which ensured a purification factor of 10.0, K of 0.2 and activity yield of 102.0%. SDS-PAGE and fibrin zymography showed that the purified protease has a molecular mass of 97kDa and an apparent isoelectric point of 5.4. When submitted to assays with different substrates and inhibitors, it showed selectivity for succinyl-l-ala-ala-pro-l-phenylalanine-p-nitroanilide and was almost completely inhibited by phenylmethylsulfonyl fluoride, behaving as a chymotrypsin-like protease. At the optimum temperature of 37°C, the enzyme residual activity was 94 and 68% of the initial one after 120 and 150min of incubation, respectively. This study demonstrated that M. subtilissimus protease has potent fibrinolytic activity compared with similar enzymes produced by solid-state fermentation, therefore it may be used as an agent for the prevention and therapy of thrombosis. Furthermore, it appears to have the advantages of low cost production and simple purification.

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Section: Determination Of the Optimal Conditions For Proteases Purifimentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Purification of a fibrinolytic protease from Mucor subtilissimus UCP 1262 by aqueous two-phase systems (PEG/sulfate)

Nascimento

Sales

Porto

et al. 2016

Journal of Chromatography B

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“…Accumulation of fibrin in the blood vessels usually increases thrombosis, leading to myocardial infarction, and other cardiovascular diseases. The blood clot fibrin, in vivo, is lysed by plasmin, which is activated from plasminogen by tissue plasminogen activator (tPA) (Venkataraman et al, 2010). The typical thrombolytic agents include urokinase (Duffy, 2002) and tPA (Collen & Lijnen, 2004).…”

Section: Introductionmentioning

confidence: 99%

A novel fibrinolytic enzymes from the Korean traditional fermented food—Jotgal: Purification and characterization

Kim¹,

Ri²,

Choe

2020

J Food Biochem

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The fibrinolytic activity in Korean traditional fermented food, Jotgal (pickled fish) was identified. Though the fibrinolytic activity could vary in different kinds of Jotgal, this activity seems to be produced by microorganisms during the natural fermentation stage. From Gonjaengijot (pickled opossum shrimp), two novel fibrinolytic enzymes named by JP‐I and JP‐II, have been purified by ethanol precipitation, Bio‐GEL P‐100 gel filtration, and DEAE‐cellulose ion‐exchange chromatography. Compared to the crude enzyme extract, the specific activity of the JP‐I and JP‐II increased 258, 85‐fold with the recovery of 22.1, 8.5%, respectively. The molecular weights of both enzymes were estimated as 36 kDa on sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS–PAGE). The optimal condition for fibrinolytic activity of JP‐I was at 50°C and pH 8.1, while that of JP‐II was at 45°C and 9.9. Both enzymes were stable at a broad range of pH (5.0 to 10.5) and have metalloprotease nature. From these results, it concludes that these enzymes could be a novel potent thrombolytic agent. Practical applications The fibrinolytic enzyme is one of the clinical agents for cardiovascular diseases which is the leading cause of morbidity and mortality worldwide with 17 million deaths every year. A variety of fibrinolytic enzymes are found and characterized from various sources such as plants, animals, and microorganisms, and new sources for fibrinolytic enzymes continue to be explored. Jotgal, widely used in Korean people's diet, is a traditional Korean seafood prepared from many different types of fishes, fish eggs, fish intestines, and shellfishes. Through an amount of research, some of fibrinolytic enzymes were found and purified from Jotgal, however, no studies have been done on fibrinolytic enzyme from opossum shrimp. In this study, the purification, enzymatic characteristics, and fibrinolytic activity of the proteases, originated from Korean traditional fermented food, Jotgal were reported. These enzymes could be novel potent thrombolytic agent.

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“…Lumbrokinase from earthworm, fibrolase from snake venom and other plasmin-like proteins directly degrade fibrin, thereby dissolving thrombi rapidly and completely (Jayalakshmi et al 2012). Although plasminogen activators and urokinase are still widely used in thrombolytic therapy, their expensive prices and undesirable side effects have prompted researchers to target novel, cheaper and safer resources (Agrebi et al 2010;Deepak et al 2010).…”

Section: Fibrinolytic Proteases As Thrombolytic Agentsmentioning

confidence: 99%

Potential application spectrum of microbial proteases for clean and green industrial production

Singh

Bajaj

2017

Energ. Ecol. Environ.

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Enzymes are the cornerstones of metabolism and constitute the fundamental basis for existence of life. However, recently enzymes are being implicated in diverse industrial processes because of their specific and fast action for efficient bioconversion of substrate to product, and their capability to save raw materials, energy and chemicals for various manufacturing processes. Enzymes are considered as environment-friendly (green) chemicals that may potentially help replacing completely or reducing the usage of hazardous chemicals for industrial processes, thus promising sustainable production and manufacturing. Among various industrial enzymes microbial proteases dominate the world enzyme market due to their multifaceted application potential in varied bioindustries like food, pharmaceutical, textile, photographic, leather and detergent. Promising applications of proteases in agricultural sector for instance may include biocontrol of pests, degumming of silk, selective delignification of hemp and wool processing. However, for successful industrial applications the proteases must be robust enough to suit the process conditions which are generally hostile. Proteases intended for industrial applications must have activity and stability over wide range of temperature and pH extremes for prolonged time periods and even in the presence of various potential enzyme inhibitors. Of various microbial proteases those from Bacillus spp. have got special significance because the latter are known for their ability to produce sturdy enzymes that might have suitability for industrial process conditions. The current article presents an interpretive summary of the recent developments on application potential of proteases for various industries.

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Medium optimization and immobilization of purified fibrinolytic URAK from Bacillus cereus NK1 on PHB nanoparticles

Cited by 35 publications

References 32 publications

Purification of a fibrinolytic protease from Mucor subtilissimus UCP 1262 by aqueous two-phase systems (PEG/sulfate)

Purification of a fibrinolytic protease from Mucor subtilissimus UCP 1262 by aqueous two-phase systems (PEG/sulfate)

A novel fibrinolytic enzymes from the Korean traditional fermented food—Jotgal: Purification and characterization

Potential application spectrum of microbial proteases for clean and green industrial production

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