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Mahiran Basri - One of the best experts on this subject based on the ideXlab platform.
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optimization of physical factors affecting the production of thermo stable Organic Solvent tolerant protease from a newly isolated halo tolerant bacillus subtilis strain rand
Microbial Cell Factories, 2009Co-Authors: Randa Abdelkareem Abusham, Raja Noor Zaliha Raja Abdul Rahman, Abu Bakar Salleh, Mahiran BasriAbstract:Many researchers have reported on the optimization of protease production; nevertheless, only a few have reported on the optimization of the production of Organic Solvent-tolerant proteases. Ironically, none has reported on thermostable Organic Solvent-tolerant protease to date. The aim of this study was to isolate the thermostable Organic Solvent-tolerant protease and identify the culture conditions which support its production. The bacteria of genus Bacillus are active producers of extra-cellular proteases, and the thermostability of enzyme production by Bacillus species has been well-studied by a number of researchers. In the present study, the Bacillus subtilis strain Rand was isolated from the contaminated soil found in Port Dickson, Malaysia. A thermostable Organic Solvent-tolerant protease producer had been identified as Bacillus subtilis strain Rand, based on the 16S rRNA analysis conducted, as well as the morphological characteristics and biochemical properties. The production of the thermostable Organic Solvent-tolerant protease was optimized by varying various physical culture conditions. Inoculation with 5.0% (v/v) of (AB600 = 0.5) inoculum size, in a culture medium (pH 7.0) and incubated for 24 h at 37°C with 200 rpm shaking, was the best culture condition which resulted in the maximum growth and production of protease (444.7 U/ml; 4042.4 U/mg). The Rand protease was not only stable in the presence of Organic Solvents, but it also exhibited a higher activity than in the absence of Organic Solvent, except for pyridine which inhibited the protease activity. The enzyme retained 100, 99 and 80% of its initial activity, after the heat treatment for 30 min at 50, 55, and 60°C, respectively. Strain Rand has been found to be able to secrete extra-cellular thermostable Organic Solvent-tolerant protease into the culture medium. The protease exhibited a remarkable stability towards temperature and Organic Solvent. This unique property makes it attractive and useful to be used in industrial applications.
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A new Organic Solvent tolerant protease from Bacillus pumilus 115b
Journal of Industrial Microbiology & Biotechnology, 2007Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Shalihah Mahamad, Mahiran BasriAbstract:Five out of the nine benzene–toulene–ethylbenzene-xylene (BTEX) tolerant bacteria that demonstrated high protease activity on skim milk agar were isolated. Among them, isolate 115b identified as Bacillus pumilus exhibited the highest protease production. The protease produced was stable in 25% (v/v) benzene and toluene and it was activated 1.7 and 2.5- fold by n -dodecane and n -tetradecane, respectively. The gene encoding the Organic Solvent tolerant protease was cloned and its nucleotide sequence determined. Sequence analysis revealed an open reading frame (ORF) of 1,149 bp that encoded a polypeptide of 383 amino acid residues. The polypeptide composed of 29 residues of signal peptide, a propeptide of 79 residues and a mature protein of 275 amino acids with a calculated molecular mass of 27,846 Da. This is the only report available to date on Organic Solvent tolerant protease from B . pumilus .
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physical factors affecting the production of Organic Solvent tolerant protease by pseudomonas aeruginosa strain k
Bioresource Technology, 2005Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Lee Poh Geok, Mahiran Basri, Abu Bakar SallehAbstract:Abstract The physical factors affecting the production of an Organic Solvent-tolerant protease from Pseudomonas aeruginosa strain K was investigated. Growth and protease production were detected from 37 to 45 °C with 37 °C being the optimum temperature for P. aeruginosa. Maximum enzyme activity was achieved at static conditions with 4.0% (v/v) inoculum. Shifting the culture from stationary to shaking condition decreased the protease production (6.0–10.0% v/v). Extracellular Organic Solvent-tolerant protease was detected over a broad pH range from 6.0 to 9.0. However, the highest yield of protease was observed at pH 7.0. Neutral media increased the protease production compared to acidic or alkaline media
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a newly isolated Organic Solvent tolerant bacillus sphaericus 205y producing Organic Solvent stable lipase
Biochemical Engineering Journal, 2003Co-Authors: Chin John Hun, Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Mahiran BasriAbstract:Abstract A total of 131 Organic Solvent tolerant isolates originating from soil samples were successfully isolated via direct plating method using 1% (v/v) of either benzene, toluene, or mixture of benzene and toluene as their sole carbon source. Six isolates demonstrated high tolerance of up to 75% (v/v) concentration of BTEX (n-benzene, toluene, ethyl benzene and p-xylene). These isolates were screened for lipolytic activity using triolein plates. Isolate 205y was found to produce the highest lipase yield in liquid medium and later identified as Bacillus sphaericus by biochemical test and 16S ribosomal DNA sequence. No work on B. sphaericus producing lipase has been reported so far. Four production media were tested and medium M3 yielded the highest lipase activity of 0.42 U ml−1 min−1 at 36 h incubation time. Stability studies in various Organic Solvents at 25% (v/v) showed that lipase from B. sphaericus 205y were not only stable but also activated by n-hexane and p-xylene by 3.5- and 2.9-folds, respectively. The activity, however, was greatly reduced in dimethyl sulfoxide and was completely inactivated by hexadecane and acetonitrile.
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Organic Solvent tolerant lipase by pseudomonas sp strain s5 stability of enzyme in Organic Solvent and physical factors affecting its production
Annals of Microbiology, 2003Co-Authors: Syarul Nataqain Baharum, Mahiran Basri, Abu Bakar Salleh, C N A Razak, M F Rahman, Raja Noor Zaliha Raja Abdul RahmanAbstract:An Organic Solvent tolerant bacterial strain was isolated. This strain was identified as Pseudomonas sp. strain S5 and shown to be BTEX (Benzene, Toluene, Ethyl-Benzene and Xylene) degrader. The strain produces a lipase that is stable in the presence of Organic Solvents such as n-hexane, cyclohexane, toluene and 1-octanol. The production of lipase by Pseudomonas sp. strain S5 was optimum at 37 °C, pH 7.0 and 6% starting inoculum.
Karl-erich Jaeger - One of the best experts on this subject based on the ideXlab platform.
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identification of Organic Solvent tolerant carboxylic ester hydrolases for Organic synthesis
Applied and Environmental Microbiology, 2020Co-Authors: Alexander Bollinger, Rebecka Molitor, Cristina Coscolin, Rainhard Koch, Karl-erich Jaeger, Stephan Thies, Manuel FerrerAbstract:Biocatalysis has emerged as an important tool in synthetic Organic chemistry enabling the chemical industry to execute reactions with high regio- or enantioselectivity and under usually mild reaction conditions while avoiding toxic waste. Target substrates and products of reactions catalyzed by carboxylic ester hydrolases are often poorly water soluble and require Organic Solvents, whereas enzymes are evolved by nature to be active in cells, i.e., in aqueous rather than Organic Solvents. Therefore, biocatalysts that withstand Organic Solvents are urgently needed. Current strategies to identify such enzymes rely on laborious tests carried out by incubation in different Organic Solvents and determination of residual activity. Here, we describe a simple assay useful for screening large libraries of carboxylic ester hydrolases for resistance and activity in water-miscible Organic Solvents. We have screened a set of 26 enzymes, most of them identified in this study, with four different water-miscible Organic Solvents. The triglyceride tributyrin was used as a substrate, and fatty acids released by enzymatic hydrolysis were detected by a pH shift indicated by the indicator dye nitrazine yellow. With this strategy, we succeeded in identifying a novel highly Organic-Solvent-tolerant esterase from Pseudomonas aestusnigri In addition, the newly identified enzymes were tested with sterically demanding substrates, which are common in pharmaceutical intermediates, and two enzymes from Alcanivorax borkumensis were identified which outcompeted the gold standard ester hydrolase CalB from Candida antarctica IMPORTANCE Major challenges hampering biotechnological applications of esterases include the requirement to accept nonnatural and chemically demanding substrates and the tolerance of the enzymes toward Organic Solvents which are often required to solubilize such substrates. We describe here a high-throughput screening strategy to identify novel Organic-Solvent-tolerant carboxylic ester hydrolases (CEs). Among these enzymes, CEs active against water-insoluble bulky substrates were identified. Our results thus contribute to fostering the identification and biotechnological application of CEs.
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Organic Solvent tolerant carboxylic ester hydrolases for Organic synthesis
Applied and Environmental Microbiology, 2020Co-Authors: Alexander Bollinger, Rebecka Molitor, Cristina Coscolin, Rainhard Koch, Karl-erich Jaeger, Stephan Thies, Manuel FerrerAbstract:ABSTRACT Biocatalysis has emerged as an important tool in synthetic Organic chemistry enabling the chemical industry to execute reactions with high regio- or enantioselectivity and under usually mild reaction conditions while avoiding toxic waste. Target substrates and products of reactions catalyzed by carboxylic ester hydrolases are often poorly water soluble and require Organic Solvents, whereas enzymes are evolved by nature to be active in cells, i.e., in aqueous rather than Organic Solvents. Therefore, biocatalysts that withstand Organic Solvents are urgently needed. Current strategies to identify such enzymes rely on laborious tests carried out by incubation in different Organic Solvents and determination of residual activity. Here, we describe a simple assay useful for screening large libraries of carboxylic ester hydrolases for resistance and activity in water-miscible Organic Solvents. We have screened a set of 26 enzymes, most of them identified in this study, with four different water-miscible Organic Solvents. The triglyceride tributyrin was used as a substrate, and fatty acids released by enzymatic hydrolysis were detected by a pH shift indicated by the indicator dye nitrazine yellow. With this strategy, we succeeded in identifying a novel highly Organic-Solvent-tolerant esterase from Pseudomonas aestusnigri. In addition, the newly identified enzymes were tested with sterically demanding substrates, which are common in pharmaceutical intermediates, and two enzymes from Alcanivorax borkumensis were identified which outcompeted the gold standard ester hydrolase CalB from Candida antarctica. IMPORTANCE Major challenges hampering biotechnological applications of esterases include the requirement to accept nonnatural and chemically demanding substrates and the tolerance of the enzymes toward Organic Solvents which are often required to solubilize such substrates. We describe here a high-throughput screening strategy to identify novel Organic-Solvent-tolerant carboxylic ester hydrolases (CEs). Among these enzymes, CEs active against water-insoluble bulky substrates were identified. Our results thus contribute to fostering the identification and biotechnological application of CEs.
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identification of Organic Solvent tolerant carboxylic ester hydrolases for Organic synthesis
Applied and Environmental Microbiology, 2020Co-Authors: Alexander Bollinger, Rebecka Molitor, Cristina Coscolin, Rainhard Koch, Karl-erich Jaeger, Stephan Thies, Manuel FerrerAbstract:Biocatalysis has emerged as an important tool in synthetic Organic chemistry enabling the chemical industry to execute reactions with high regio- or enantioselectivity and under usually mild reaction conditions while avoiding toxic waste. Target substrates and products of reactions catalyzed by carboxylic ester hydrolases are often poorly water soluble and require Organic Solvents, whereas enzymes are evolved by nature to be active in cells, i.e., in aqueous rather than Organic Solvents. Therefore, biocatalysts that withstand Organic Solvents are urgently needed. Current strategies to identify such enzymes rely on laborious tests carried out by incubation in different Organic Solvents and determination of residual activity. Here, we describe a simple assay useful for screening large libraries of carboxylic ester hydrolases for resistance and activity in water-miscible Organic Solvents. We have screened a set of 26 enzymes, most of them identified in this study, with four different water-miscible Organic Solvents. The triglyceride tributyrin was used as a substrate, and fatty acids released by enzymatic hydrolysis were detected by a pH shift indicated by the indicator dye nitrazine yellow. With this strategy, we succeeded in identifying a novel highly Organic-Solvent-tolerant esterase from Pseudomonas aestusnigri In addition, the newly identified enzymes were tested with sterically demanding substrates, which are common in pharmaceutical intermediates, and two enzymes from Alcanivorax borkumensis were identified which outcompeted the gold standard ester hydrolase CalB from Candida antarctica IMPORTANCE Major challenges hampering biotechnological applications of esterases include the requirement to accept nonnatural and chemically demanding substrates and the tolerance of the enzymes toward Organic Solvents which are often required to solubilize such substrates. We describe here a high-throughput screening strategy to identify novel Organic-Solvent-tolerant carboxylic ester hydrolases (CEs). Among these enzymes, CEs active against water-insoluble bulky substrates were identified. Our results thus contribute to fostering the identification and biotechnological application of CEs.
Abu Bakar Salleh - One of the best experts on this subject based on the ideXlab platform.
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optimization of physical factors affecting the production of thermo stable Organic Solvent tolerant protease from a newly isolated halo tolerant bacillus subtilis strain rand
Microbial Cell Factories, 2009Co-Authors: Randa Abdelkareem Abusham, Raja Noor Zaliha Raja Abdul Rahman, Abu Bakar Salleh, Mahiran BasriAbstract:Many researchers have reported on the optimization of protease production; nevertheless, only a few have reported on the optimization of the production of Organic Solvent-tolerant proteases. Ironically, none has reported on thermostable Organic Solvent-tolerant protease to date. The aim of this study was to isolate the thermostable Organic Solvent-tolerant protease and identify the culture conditions which support its production. The bacteria of genus Bacillus are active producers of extra-cellular proteases, and the thermostability of enzyme production by Bacillus species has been well-studied by a number of researchers. In the present study, the Bacillus subtilis strain Rand was isolated from the contaminated soil found in Port Dickson, Malaysia. A thermostable Organic Solvent-tolerant protease producer had been identified as Bacillus subtilis strain Rand, based on the 16S rRNA analysis conducted, as well as the morphological characteristics and biochemical properties. The production of the thermostable Organic Solvent-tolerant protease was optimized by varying various physical culture conditions. Inoculation with 5.0% (v/v) of (AB600 = 0.5) inoculum size, in a culture medium (pH 7.0) and incubated for 24 h at 37°C with 200 rpm shaking, was the best culture condition which resulted in the maximum growth and production of protease (444.7 U/ml; 4042.4 U/mg). The Rand protease was not only stable in the presence of Organic Solvents, but it also exhibited a higher activity than in the absence of Organic Solvent, except for pyridine which inhibited the protease activity. The enzyme retained 100, 99 and 80% of its initial activity, after the heat treatment for 30 min at 50, 55, and 60°C, respectively. Strain Rand has been found to be able to secrete extra-cellular thermostable Organic Solvent-tolerant protease into the culture medium. The protease exhibited a remarkable stability towards temperature and Organic Solvent. This unique property makes it attractive and useful to be used in industrial applications.
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A new Organic Solvent tolerant protease from Bacillus pumilus 115b
Journal of Industrial Microbiology & Biotechnology, 2007Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Shalihah Mahamad, Mahiran BasriAbstract:Five out of the nine benzene–toulene–ethylbenzene-xylene (BTEX) tolerant bacteria that demonstrated high protease activity on skim milk agar were isolated. Among them, isolate 115b identified as Bacillus pumilus exhibited the highest protease production. The protease produced was stable in 25% (v/v) benzene and toluene and it was activated 1.7 and 2.5- fold by n -dodecane and n -tetradecane, respectively. The gene encoding the Organic Solvent tolerant protease was cloned and its nucleotide sequence determined. Sequence analysis revealed an open reading frame (ORF) of 1,149 bp that encoded a polypeptide of 383 amino acid residues. The polypeptide composed of 29 residues of signal peptide, a propeptide of 79 residues and a mature protein of 275 amino acids with a calculated molecular mass of 27,846 Da. This is the only report available to date on Organic Solvent tolerant protease from B . pumilus .
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physical factors affecting the production of Organic Solvent tolerant protease by pseudomonas aeruginosa strain k
Bioresource Technology, 2005Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Lee Poh Geok, Mahiran Basri, Abu Bakar SallehAbstract:Abstract The physical factors affecting the production of an Organic Solvent-tolerant protease from Pseudomonas aeruginosa strain K was investigated. Growth and protease production were detected from 37 to 45 °C with 37 °C being the optimum temperature for P. aeruginosa. Maximum enzyme activity was achieved at static conditions with 4.0% (v/v) inoculum. Shifting the culture from stationary to shaking condition decreased the protease production (6.0–10.0% v/v). Extracellular Organic Solvent-tolerant protease was detected over a broad pH range from 6.0 to 9.0. However, the highest yield of protease was observed at pH 7.0. Neutral media increased the protease production compared to acidic or alkaline media
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a newly isolated Organic Solvent tolerant bacillus sphaericus 205y producing Organic Solvent stable lipase
Biochemical Engineering Journal, 2003Co-Authors: Chin John Hun, Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Mahiran BasriAbstract:Abstract A total of 131 Organic Solvent tolerant isolates originating from soil samples were successfully isolated via direct plating method using 1% (v/v) of either benzene, toluene, or mixture of benzene and toluene as their sole carbon source. Six isolates demonstrated high tolerance of up to 75% (v/v) concentration of BTEX (n-benzene, toluene, ethyl benzene and p-xylene). These isolates were screened for lipolytic activity using triolein plates. Isolate 205y was found to produce the highest lipase yield in liquid medium and later identified as Bacillus sphaericus by biochemical test and 16S ribosomal DNA sequence. No work on B. sphaericus producing lipase has been reported so far. Four production media were tested and medium M3 yielded the highest lipase activity of 0.42 U ml−1 min−1 at 36 h incubation time. Stability studies in various Organic Solvents at 25% (v/v) showed that lipase from B. sphaericus 205y were not only stable but also activated by n-hexane and p-xylene by 3.5- and 2.9-folds, respectively. The activity, however, was greatly reduced in dimethyl sulfoxide and was completely inactivated by hexadecane and acetonitrile.
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Organic Solvent tolerant lipase by pseudomonas sp strain s5 stability of enzyme in Organic Solvent and physical factors affecting its production
Annals of Microbiology, 2003Co-Authors: Syarul Nataqain Baharum, Mahiran Basri, Abu Bakar Salleh, C N A Razak, M F Rahman, Raja Noor Zaliha Raja Abdul RahmanAbstract:An Organic Solvent tolerant bacterial strain was isolated. This strain was identified as Pseudomonas sp. strain S5 and shown to be BTEX (Benzene, Toluene, Ethyl-Benzene and Xylene) degrader. The strain produces a lipase that is stable in the presence of Organic Solvents such as n-hexane, cyclohexane, toluene and 1-octanol. The production of lipase by Pseudomonas sp. strain S5 was optimum at 37 °C, pH 7.0 and 6% starting inoculum.
Hiroyasu Ogino - One of the best experts on this subject based on the ideXlab platform.
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Organic Solvent-tolerant enzymes
Biochemical Engineering Journal, 2010Co-Authors: Noriyuki Doukyu, Hiroyasu OginoAbstract:Abstract The use of Organic Solvents as reaction media for enzymatic reactions provides numerous industrially attractive advantages compared to traditional aqueous reaction systems. Despite the advantages, native enzymes almost universally exhibit low activities and/or stabilities in the presence of Organic Solvents. This inactivation of the enzymes by Organic Solvents results in significant limitation of the enzymatic reaction process. Numerous attempts have been made to improve enzyme activity and stability in the presence of Organic Solvents using methods based on protein engineering and chemical or physical modification. Most enzymes used in these studies did not originally exhibit high activity and stability in the presence of Organic Solvents because they were not screened as Organic Solvent-tolerant enzymes. Recently, various attempts have been made to screen enzymes that naturally possess Organic Solvent-tolerance form various microorganisms including Organic Solvent-tolerant bacteria, thermophiles, halophiles and mesophiles. These Organic Solvent-tolerant enzymes are expected to have potential for applications in industrial chemical processes.
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enhancement of the Organic Solvent stability of the lst 03 lipase by directed evolution
Biotechnology Progress, 2009Co-Authors: Takuya Kawata, Hiroyasu OginoAbstract:LST-03 lipase from an Organic Solvent-tolerant Pseudomonas aeruginosa LST-03 has high stability and activity in the presence of various Organic Solvents. In this research, enhancement of Organic Solvent-stability of LST-03 lipase was attempted by directed evolution. The structural gene of the LST-03 lipase was amplified by the error prone-PCR method. Organic Solvent-stability of the mutated lipases was assayed by formation of a clear zone of agar which contained dimethyl sulfoxide (DMSO) and tri-n-butyrin and which overlaid a plate medium. And the Organic Solvent-stability was also confirmed by measuring the half-life of activity in the presence of DMSO. Four mutated enzymes were selected on the basis of their high Organic Solvent-stability in the presence of DMSO. The Organic Solvent-stabilities of mutated LST-03 lipase in the presence of various Organic Solvents were measured and their mutated amino acid residues were identified. The half-lives of the LST-03-R65 lipase in the presence of cyclohexane and n-decane were about 9 to 11-fold longer than those of the wild-type lipase, respectively. Some substituted amino acid residues of mutated LST-03 lipases have been located at the surface of the enzyme molecules, while some other amino acid residues have been changed from neutral to basic residues. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009
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role of intermolecular disulfide bonds of the Organic Solvent stable pst 01 protease in its Organic Solvent stability
Applied and Environmental Microbiology, 2001Co-Authors: Hiroyasu Ogino, Takeshi Uchiho, Jyunko Yokoo, Reina Kobayashi, Rikiya Ichise, Haruo IshikawaAbstract:The PST-01 protease is secreted by the Organic Solvent-tolerant microorganism Pseudomonas aeruginosa PST-01 and is stable in the presence of various Organic Solvents. Therefore, the PST-01 strain and the PST-01 protease are very useful for fermentation and reactions in the presence of Organic Solvents, respectively. The Organic Solvent-stable PST-01 protease has two disulfide bonds (between Cys-30 and Cys-58 and between Cys-270 and Cys-297) in its molecule. Mutant PST-01 proteases in which one or both of the disulfide bonds were deleted were constructed by site-directed mutagenesis, and the effect of the disulfide bonds on the activity and the various stabilities was investigated. The disulfide bond between Cys-270 and Cys-297 in the PST-01 protease was found to be essential for its activity. The disulfide bond between Cys-30 and Cys-58 played an important role in the Organic Solvent stability of the PST-01 protease.
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purification and characterization of Organic Solvent stable lipase from Organic Solvent tolerant pseudomonas aeruginosa lst 03
Journal of Bioscience and Bioengineering, 1999Co-Authors: Hiroyasu Ogino, Satoshi Nakagawa, Kaori Shinya, Toshiaki Muto, Nobuyuki Fujimura, Masahiro Yasuda, Haruo IshikawaAbstract:An Organic Solvent-stable lipase (LST-03 lipase) secreted into the culture broth of the Organic Solvent-tolerant Pseudomonas aeruginosa LST-03 was purified by ion-exchange and hydrophobic interaction chromatography in the presence of 2-propanol. The purified enzyme was homogeneous as determined by SDS-PAGE. The molecular mass of the lipase was estimated to be 27.1 kDa by SDS-PAGE and 36 kDa by gel filtration. The optimum pH and temperature were 6.0 and 37°C. LST-03 lipase was stable at pH 5–8 and below 40°C. Its hydrolytic activity was highest against tricaproin (C6), methyl octanoate (C8), and coconut oil respectively among the triacylglycerols, fatty acid methyl esters, and natural oils investigated. The enzyme cleaved not only the 1,3-positioned ester bonds, but also the 2-positioned ester bond of triolein. It exhibited high levels of activity in the presence of n-decane, n-octane, DMSO, and DMF as well as in the absence of an Organic Solvent. In addition, LST-03 lipase was stabler in the presence of n-decane, ethyleneglycol, DMSO, n-octane, n-heptane, isooctane, and cyclohexane than in the absence of an Organic Solvent.
Raja Noor Zaliha Raja Abd Rahman - One of the best experts on this subject based on the ideXlab platform.
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A new Organic Solvent tolerant protease from Bacillus pumilus 115b
Journal of Industrial Microbiology & Biotechnology, 2007Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Shalihah Mahamad, Mahiran BasriAbstract:Five out of the nine benzene–toulene–ethylbenzene-xylene (BTEX) tolerant bacteria that demonstrated high protease activity on skim milk agar were isolated. Among them, isolate 115b identified as Bacillus pumilus exhibited the highest protease production. The protease produced was stable in 25% (v/v) benzene and toluene and it was activated 1.7 and 2.5- fold by n -dodecane and n -tetradecane, respectively. The gene encoding the Organic Solvent tolerant protease was cloned and its nucleotide sequence determined. Sequence analysis revealed an open reading frame (ORF) of 1,149 bp that encoded a polypeptide of 383 amino acid residues. The polypeptide composed of 29 residues of signal peptide, a propeptide of 79 residues and a mature protein of 275 amino acids with a calculated molecular mass of 27,846 Da. This is the only report available to date on Organic Solvent tolerant protease from B . pumilus .
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physical factors affecting the production of Organic Solvent tolerant protease by pseudomonas aeruginosa strain k
Bioresource Technology, 2005Co-Authors: Raja Noor Zaliha Raja Abd Rahman, Lee Poh Geok, Mahiran Basri, Abu Bakar SallehAbstract:Abstract The physical factors affecting the production of an Organic Solvent-tolerant protease from Pseudomonas aeruginosa strain K was investigated. Growth and protease production were detected from 37 to 45 °C with 37 °C being the optimum temperature for P. aeruginosa. Maximum enzyme activity was achieved at static conditions with 4.0% (v/v) inoculum. Shifting the culture from stationary to shaking condition decreased the protease production (6.0–10.0% v/v). Extracellular Organic Solvent-tolerant protease was detected over a broad pH range from 6.0 to 9.0. However, the highest yield of protease was observed at pH 7.0. Neutral media increased the protease production compared to acidic or alkaline media
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a newly isolated Organic Solvent tolerant bacillus sphaericus 205y producing Organic Solvent stable lipase
Biochemical Engineering Journal, 2003Co-Authors: Chin John Hun, Raja Noor Zaliha Raja Abd Rahman, Abu Bakar Salleh, Mahiran BasriAbstract:Abstract A total of 131 Organic Solvent tolerant isolates originating from soil samples were successfully isolated via direct plating method using 1% (v/v) of either benzene, toluene, or mixture of benzene and toluene as their sole carbon source. Six isolates demonstrated high tolerance of up to 75% (v/v) concentration of BTEX (n-benzene, toluene, ethyl benzene and p-xylene). These isolates were screened for lipolytic activity using triolein plates. Isolate 205y was found to produce the highest lipase yield in liquid medium and later identified as Bacillus sphaericus by biochemical test and 16S ribosomal DNA sequence. No work on B. sphaericus producing lipase has been reported so far. Four production media were tested and medium M3 yielded the highest lipase activity of 0.42 U ml−1 min−1 at 36 h incubation time. Stability studies in various Organic Solvents at 25% (v/v) showed that lipase from B. sphaericus 205y were not only stable but also activated by n-hexane and p-xylene by 3.5- and 2.9-folds, respectively. The activity, however, was greatly reduced in dimethyl sulfoxide and was completely inactivated by hexadecane and acetonitrile.
