Microbial alkaline proteases

Kumar, C. Ganesh; Takagi, Hiroshi

doi:10.1016/s0734-9750(99)00027-0

Cited by 649 publications

(159 citation statements)

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“…Extracellular proteases are important for the hydrolysis of proteins in cell-free environments and enable the cell to absorb and utilize hydrolytic products 4 . At the same time, these extracellular proteases have also been commercially exploited to assist protein degradation in various industrial processes 5,6 . It is important to screen the media components and fermentation conditions for cost-efficient enzyme production; proteases are generally produced by submerged fermentation.…”

Section: Introductionmentioning

confidence: 99%

Screening of Nutritional Parameters for the Production of Protease from Aspergillus Oryzae

Gedela

Lokeswari

Jayaraju

2006

Journal of Chemistry

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Production of protease enzyme by fungus Aspergillus oryzae was investigated. The proteolytic activity was observed when the fungus was grown in the medium containing glucose, malt extract, yeast extract, peptone, K 2 HPO 4 , MgSO 4 and FeSO 4 . The present paper describes the screening of media components and fermentation conditions in shake flask. The organism utilized carbon sources glucose, fructose, sucrose, lactose, dextrin and starch among them glucose was found to be the best carbon source, for nitrogen sources various inorganic and organic media components were investigated among them peptone is found to be the best nitrogen source. 1% cottonseed followed by 2% Soya bean meal was found to be the best inducer. With optimized media two-fold increase in the protease production. The fungus growth depends on the concentration of carbon, nitrogen and salt solution, where as the enzyme production was also influenced by the culture time, pH and interaction between these two variables.

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Section: Introductionmentioning

confidence: 99%

Screening of Nutritional Parameters for the Production of Protease from Aspergillus Oryzae

Gedela

Lokeswari

Jayaraju

2006

Journal of Chemistry

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“…Crude preparations are generally more stable than the purified enzymes. Therefore storage requirements of the purified enzymes are of the prime concern to enzyme manufacturers (35). Majority of the proteins exhibit decreased stability when are not in their native environment.…”

Section: Discussionmentioning

confidence: 99%

Optimization of Parameters that Affect the Activity of the Alkaline Protease from Halotolerant Bacterium, Bacillus acquimaris VITP4, by the Application of Response Surface Methodology and Evaluation of the Storage Stability of the Enzyme

2016

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A great deal of efforts is focused on meeting the increasing demand for novel microbial catalysts that are capable of functioning under extreme conditions, and therefore, the production of these novel microbial enzymes with appreciable activity and stability in the harsh environments (1). Enzymes from organisms which can grow in 0-15% of the NaCl will have great potential for the use in the industries because of their inherent ability to be active and stable both in the presence as well as in the absence of salt. However, most of these enzymes require metal ions for the maintenance of the structure and/or stability (2). These enzymes are active and stable under conditions that cause in a lower activity, which include high salt concentrations or organic solvents (3). In most of the cases, measurable proteolytic activity is achieved upon supplementation of cofactors like divalent metal ions in the assaying buffer. Also loss of enzymatic activity can be restored by incubating the enzyme with excess metal ions (4). Background: It was previously shown that the activity of a serine protease from a moderately halotolerant Bacillus aquimaris VITP4 strain is active in a wide range of pH and temperatures and could be modulated by the presence of the divalent metal ions. Objectives: In the present study, a quantitative analysis was done in order to explore the parameters that are contributing to the protease activity. Materials and Methods: Changes in the secondary structure of the enzyme was determined by circular dichroism analysis. The conditions for the optimal activity was investigated by Response Surface Methodology. Stability of the enzyme was determined by thermal inactivation experiments. Results:The initial one-factor-at-a-time experiments have indicated that the activity of the enzyme could be enhanced not only by the presence of low concentrations of NaCl but also by divalent metal ions, such as Ca 2+ , Mn 2+ and Cu 2+ . A clear dependence of the activity to the secondary structure of the enzyme could be established using circular dichroism spectroscopy. In the next level of optimization, four factors; viz. pH, temperature, concentration of Ca 2+ , and Mn 2+ were used to optimize the conditions required for the maximal activity of the enzyme by Response Surface Methodology, and the data could be explained using quadratic model. Under optimal condition of 43°C, pH 8.0, 8.2 mM Ca 2+ , and 4.3 mM Mn 2+ a 1.5 times enhancement in the enzyme activity could be achieved. The storage stability of the enzyme under these selected conditions has indicated a non-linear relation between the conditions for the enzymatic activity as well as stability. However, the condition for the maximal stability (267±18 min) has corresponded to that of the optimal conditions for the maximal activity. Conclusions: This study, for the first time, has explored the possibility of using statistical methods for identifying the optimal conditions for alkaline protease activity isolated from the halotolerant Bacillus aquimaris VITP4.

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“…Proteases obtained from the microbial community are preferred for the large-scale production of proteases due to their fast growth and simple life cycle for the generation of new recombinant enzymes with desired properties (Kumar and Takagi, 1999;Jisha et al, 2013). Achromobacter protease I (API) is a lysine-specific serine protease that specifically hydrolyzes the lysyl peptide bond (Ohara et al, 1989).…”

Section: Introductionmentioning

confidence: 99%

Genetic markers for detection of Escherichia coli K-12 harboring ampicillin-resistance plasmid from an industrial wastewater treatment effluent pond

et al. 2016

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ABSTRACT. Biotechnology industries that use recombinant DNA technology are potential sources for release of genetically modified organisms to the environment. Antibiotic-resistance marker genes are commonly used for recombinant bacteria selection. One example is the marker gene coding for β-lactamase (bla) in plasmids found in Escherichia coli K-12. The aim of this study was to provide an approach to develop a molecular method for genetic marker detection in E. coli K-12 harboring bla genes from an industrial wastewater treatment effluent pond (IWTEP). For the detection of bla and Achromobacter lyticus protease I (api) genes in samples from IWTEP, we employed multiplex polymerase chain reaction (PCR) using E. coli K-12 genetic marker detection primers, previously described in the literature, and primers designed in our laboratory. The microbiological screening method resulted in 22 bacterial colony-forming units isolated from three different IWTEP harvesting points. The multiplex PCR amplicons showed that five isolates were positive for the bla gene marker and negative for the E. coli K-12 and api genes. The 16S rRNA regions of positive microorganisms carrying the bla gene were genotyped by the MicroSeq ® 500 system. The bacteria found were Escherichia spp (3/5), Chromobacterium spp (1/5), and Aeromonas spp (1/5). None of the 22 isolated microorganisms presented the molecular pattern of E. coli K-12 harboring the bla gene. The presence of microorganisms positive for the bla gene and negative for E. coli K-12 harboring bla genes at IWTEP suggests that the ampicillin resistance found in the isolated bacteria could be from microorganisms other than the E. coli K-12 strain harboring plasmid.

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Microbial alkaline proteases

Cited by 649 publications

References 211 publications

Screening of Nutritional Parameters for the Production of Protease from Aspergillus Oryzae

Screening of Nutritional Parameters for the Production of Protease from Aspergillus Oryzae

Optimization of Parameters that Affect the Activity of the Alkaline Protease from Halotolerant Bacterium, Bacillus acquimaris VITP4, by the Application of Response Surface Methodology and Evaluation of the Storage Stability of the Enzyme

Genetic markers for detection of Escherichia coli K-12 harboring ampicillin-resistance plasmid from an industrial wastewater treatment effluent pond

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