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Chit Laa Poh - One of the best experts on this subject based on the ideXlab platform.
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characterization of hbze encoded gentisate 1 2 dioxygenase from Pseudomonas Alcaligenes ncimb 9867
Research in Microbiology, 2007Co-Authors: Chew Chieng Yeo, Chew Ling Tan, Xiaoli Gao, Bing Zhao, Chit Laa PohAbstract:Abstract Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) is known to synthesize two isofunctional gentisate 1,2-dioxygenases (GDO; EC 1.13.11.4) as well as other enzymes involved in the degradation of xylenols and cresols via the gentisate pathway. The hbzE gene encoding what is possibly the strictly inducible gentisate 1,2-dioxygenase II (GDO-II) was cloned, overexpressed and purified as a hexahistidine fusion protein from Escherichia coli . Active recombinant GDO-II had an estimated molecular mass of 150 kDa and is likely a tetrameric protein with a subunit mass of ∼40 kDa, similar to the previously characterized gentisate 1,2-dioxygenase I (GDO-I) encoded by xlnE . However, GDO-II was unable to utilize gentisate that is substituted at the carbon-4 position, unlike GDO-I which had broader substrate specificity. GDO-II also possessed different kinetic characteristics when compared to GDO-I. The hbzE -encoded GDO-II shared higher sequence identities (53%) with GDOs from Ralstonia sp. U2 and Polaromonas naphthalenivorans CJ2, compared with only 35% identity with the xlnE -encoded GDO-I. The hbzE gene was found to be part of a cluster of nine genes including the putative regulatory gene designated hbzR , which encodes an LysR-type regulator and is divergently transcribed from the other genes of the hbzHIJKLFED cluster.
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proteome analysis of heat shock protein expression in Pseudomonas Alcaligenes ncimb 9867 in response to gentisate exposure and elevated growth temperature
Biotechnology and Bioengineering, 2007Co-Authors: Chew Chieng Yeo, Chew Ling Tan, Bing Zhao, Chit Laa PohAbstract:Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) degrades xylenols and cresols via the gentisate pathway. P25X expresses two isofunctional gentisate 1,2-dioxygenases (GDO I and GDO II). The expression of both GDOs was not detected when P25X cells were grown at 42°C, even in the presence of gentisate. A total of 19 heat shock proteins (Hsps) belonging to the Hsp100, Hsp90, Hsp70, Hsp60, Hsp45, and small heat shock protein (sHsp) families were identified among the protein spots that were either newly detected or were expressed at levels of at least twofold higher when P25X cells were cultured at 32 or 42°C in the presence and absence of gentisate. Among these, 16 Hsps were commonly expressed at 42°C. Two additional Hsps (H5 and H13) from the Hsp90 and Hsp60 families, respectively, were expressed only when P25X cells were grown at 42°C and in the presence of gentisate. A protein of the sHsp (H16) family was expressed only in the presence of gentisate at 32°C but not at 42°C. The GroEL chaperonins of the Hsp60 family comprised the largest group of Hsps identified and exhibited high level of expression at 42°C following gentisate exposure. Biotechnol. Bioeng. 2007;97: 506–514. © 2006 Wiley Periodicals, Inc.
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molecular and biochemical characterization of the xlnd encoded 3 hydroxybenzoate 6 hydroxylase involved in the degradation of 2 5 xylenol via the gentisate pathway in Pseudomonas Alcaligenes ncimb 9867
Journal of Bacteriology, 2005Co-Authors: Xiaoli Gao, Chew Chieng Yeo, Chit Laa Poh, Chew Ling TanAbstract:The xlnD gene from Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) was shown to encode 3-hydroxybenzoate 6-hydroxylase I, the enzyme that catalyzes the NADH-dependent conversion of 3-hydroxybenzoate to gentisate. Active recombinant XlnD was purified as a hexahistidine fusion protein from Escherichia coli, had an estimated molecular mass of 130 kDa, and is probably a trimeric protein with a subunit mass of 43 kDa. This is in contrast to the monomeric nature of the few 3-hydroxybenzoate 6-hydroxylases that have been characterized thus far. Like other 3-hydroxybenzoate 6-hydroxylases, XlnD could utilize either NADH or NADPH as the electron donor. P25X harbors a second 3-hydroxybenzoate 6-hydroxylase II that was strictly inducible by specific aromatic substrates. However, the degradation of 2,5-xylenol and 3,5-xylenol in strain P25X was found to be dependent on the xlnD-encoded 6-hydroxylase I and not the second, strictly inducible 6-hydroxylase II.
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replacement of tyrosine 181 by phenylalanine in gentisate 1 2 dioxygenase i from Pseudomonas Alcaligenes ncimb 9867 enhances catalytic activities
Journal of Bacteriology, 2005Co-Authors: Chew Ling Tan, Chew Chieng Yeo, Chit Laa Poh, H E KhooAbstract:xlnE, encoding gentisate 1,2-dioxygenase (EC 1.13.11.4), from Pseudomonas Alcaligenes (P25X) was mutagenized by site-directed mutagenesis. The mutant enzyme, Y181F, demonstrated 4-, 3-, 6-, and 16-fold increases in relative activity towards gentisate and 3-fluoro-, 4-methyl-, and 3-methylgentisate, respectively. The specific mutation conferred a 13-fold higher catalytic efficiency (kcat/Km) on Y181F towards 3-methylgentisate than that of the wild-type enzyme.
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proteome investigation of the global regulatory role of σ54 in response to gentisate induction in Pseudomonas Alcaligenes ncimb 9867
Proteomics, 2005Co-Authors: Bing Zhao, Chew Chieng Yeo, Chit Laa PohAbstract:Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) utilizes the gentisate pathway for the degradation of aromatic hydrocarbons. The gene encoding the alternative sigma (sigma) factor sigma(54), rpoN, was cloned from strain P25X and a rpoN knock-out strain, designated G54, was constructed by insertional inactivation with a kanamycin resistance gene cassette. The role of sigma(54) in the physiological response of P. Alcaligenes P25X to gentisate induction was assessed by comparing the global protein expression profiles of the wild-type P25X with the rpoN mutant strain G54. Analysis of two-dimensional polyacrylamide gel electrophoresis gels showed that 39 out of 355 prominent protein spots exhibited differential expression as a result of the insertional inactivation of rpoN. Identification of the protein spots by matrix-assisted laser desorption/ionization-time of flight/time of flight revealed a wide diversity of proteins that are affected by the sigma(54) mutation, the largest group being proteins that are involved in carbon metabolism. The strictly inducible gentisate 1,2-dioxygenase, one of two isofunctional copies of the key enzyme in the gentisate pathway, and enzymes of the TCA cycle, pyruvate metabolism and gluconeogenesis were part of this group. Other proteins that are part of the sigma(54) regulon include enzymes implicated in nitrogen metabolism, transport proteins, stress-response proteins and proteins involved in cell motility. The results of this study showed that sigma(54) plays a global regulatory role in the expression of a wide variety of genes in P. Alcaligenes, including the wild-type response to the presence of the aromatic inducer, gentisate.
Chew Chieng Yeo - One of the best experts on this subject based on the ideXlab platform.
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characterization of hbze encoded gentisate 1 2 dioxygenase from Pseudomonas Alcaligenes ncimb 9867
Research in Microbiology, 2007Co-Authors: Chew Chieng Yeo, Chew Ling Tan, Xiaoli Gao, Bing Zhao, Chit Laa PohAbstract:Abstract Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) is known to synthesize two isofunctional gentisate 1,2-dioxygenases (GDO; EC 1.13.11.4) as well as other enzymes involved in the degradation of xylenols and cresols via the gentisate pathway. The hbzE gene encoding what is possibly the strictly inducible gentisate 1,2-dioxygenase II (GDO-II) was cloned, overexpressed and purified as a hexahistidine fusion protein from Escherichia coli . Active recombinant GDO-II had an estimated molecular mass of 150 kDa and is likely a tetrameric protein with a subunit mass of ∼40 kDa, similar to the previously characterized gentisate 1,2-dioxygenase I (GDO-I) encoded by xlnE . However, GDO-II was unable to utilize gentisate that is substituted at the carbon-4 position, unlike GDO-I which had broader substrate specificity. GDO-II also possessed different kinetic characteristics when compared to GDO-I. The hbzE -encoded GDO-II shared higher sequence identities (53%) with GDOs from Ralstonia sp. U2 and Polaromonas naphthalenivorans CJ2, compared with only 35% identity with the xlnE -encoded GDO-I. The hbzE gene was found to be part of a cluster of nine genes including the putative regulatory gene designated hbzR , which encodes an LysR-type regulator and is divergently transcribed from the other genes of the hbzHIJKLFED cluster.
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proteome analysis of heat shock protein expression in Pseudomonas Alcaligenes ncimb 9867 in response to gentisate exposure and elevated growth temperature
Biotechnology and Bioengineering, 2007Co-Authors: Chew Chieng Yeo, Chew Ling Tan, Bing Zhao, Chit Laa PohAbstract:Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) degrades xylenols and cresols via the gentisate pathway. P25X expresses two isofunctional gentisate 1,2-dioxygenases (GDO I and GDO II). The expression of both GDOs was not detected when P25X cells were grown at 42°C, even in the presence of gentisate. A total of 19 heat shock proteins (Hsps) belonging to the Hsp100, Hsp90, Hsp70, Hsp60, Hsp45, and small heat shock protein (sHsp) families were identified among the protein spots that were either newly detected or were expressed at levels of at least twofold higher when P25X cells were cultured at 32 or 42°C in the presence and absence of gentisate. Among these, 16 Hsps were commonly expressed at 42°C. Two additional Hsps (H5 and H13) from the Hsp90 and Hsp60 families, respectively, were expressed only when P25X cells were grown at 42°C and in the presence of gentisate. A protein of the sHsp (H16) family was expressed only in the presence of gentisate at 32°C but not at 42°C. The GroEL chaperonins of the Hsp60 family comprised the largest group of Hsps identified and exhibited high level of expression at 42°C following gentisate exposure. Biotechnol. Bioeng. 2007;97: 506–514. © 2006 Wiley Periodicals, Inc.
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molecular and biochemical characterization of the xlnd encoded 3 hydroxybenzoate 6 hydroxylase involved in the degradation of 2 5 xylenol via the gentisate pathway in Pseudomonas Alcaligenes ncimb 9867
Journal of Bacteriology, 2005Co-Authors: Xiaoli Gao, Chew Chieng Yeo, Chit Laa Poh, Chew Ling TanAbstract:The xlnD gene from Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) was shown to encode 3-hydroxybenzoate 6-hydroxylase I, the enzyme that catalyzes the NADH-dependent conversion of 3-hydroxybenzoate to gentisate. Active recombinant XlnD was purified as a hexahistidine fusion protein from Escherichia coli, had an estimated molecular mass of 130 kDa, and is probably a trimeric protein with a subunit mass of 43 kDa. This is in contrast to the monomeric nature of the few 3-hydroxybenzoate 6-hydroxylases that have been characterized thus far. Like other 3-hydroxybenzoate 6-hydroxylases, XlnD could utilize either NADH or NADPH as the electron donor. P25X harbors a second 3-hydroxybenzoate 6-hydroxylase II that was strictly inducible by specific aromatic substrates. However, the degradation of 2,5-xylenol and 3,5-xylenol in strain P25X was found to be dependent on the xlnD-encoded 6-hydroxylase I and not the second, strictly inducible 6-hydroxylase II.
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replacement of tyrosine 181 by phenylalanine in gentisate 1 2 dioxygenase i from Pseudomonas Alcaligenes ncimb 9867 enhances catalytic activities
Journal of Bacteriology, 2005Co-Authors: Chew Ling Tan, Chew Chieng Yeo, Chit Laa Poh, H E KhooAbstract:xlnE, encoding gentisate 1,2-dioxygenase (EC 1.13.11.4), from Pseudomonas Alcaligenes (P25X) was mutagenized by site-directed mutagenesis. The mutant enzyme, Y181F, demonstrated 4-, 3-, 6-, and 16-fold increases in relative activity towards gentisate and 3-fluoro-, 4-methyl-, and 3-methylgentisate, respectively. The specific mutation conferred a 13-fold higher catalytic efficiency (kcat/Km) on Y181F towards 3-methylgentisate than that of the wild-type enzyme.
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proteome investigation of the global regulatory role of σ54 in response to gentisate induction in Pseudomonas Alcaligenes ncimb 9867
Proteomics, 2005Co-Authors: Bing Zhao, Chew Chieng Yeo, Chit Laa PohAbstract:Pseudomonas Alcaligenes NCIMB 9867 (strain P25X) utilizes the gentisate pathway for the degradation of aromatic hydrocarbons. The gene encoding the alternative sigma (sigma) factor sigma(54), rpoN, was cloned from strain P25X and a rpoN knock-out strain, designated G54, was constructed by insertional inactivation with a kanamycin resistance gene cassette. The role of sigma(54) in the physiological response of P. Alcaligenes P25X to gentisate induction was assessed by comparing the global protein expression profiles of the wild-type P25X with the rpoN mutant strain G54. Analysis of two-dimensional polyacrylamide gel electrophoresis gels showed that 39 out of 355 prominent protein spots exhibited differential expression as a result of the insertional inactivation of rpoN. Identification of the protein spots by matrix-assisted laser desorption/ionization-time of flight/time of flight revealed a wide diversity of proteins that are affected by the sigma(54) mutation, the largest group being proteins that are involved in carbon metabolism. The strictly inducible gentisate 1,2-dioxygenase, one of two isofunctional copies of the key enzyme in the gentisate pathway, and enzymes of the TCA cycle, pyruvate metabolism and gluconeogenesis were part of this group. Other proteins that are part of the sigma(54) regulon include enzymes implicated in nitrogen metabolism, transport proteins, stress-response proteins and proteins involved in cell motility. The results of this study showed that sigma(54) plays a global regulatory role in the expression of a wide variety of genes in P. Alcaligenes, including the wild-type response to the presence of the aromatic inducer, gentisate.
Lirong Yang - One of the best experts on this subject based on the ideXlab platform.
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improving Pseudomonas Alcaligenes lipase s diastereopreference in hydrolysis of diastereomeric mixture of menthyl propionate by site directed mutagenesis
Biotechnology and Bioprocess Engineering, 2014Co-Authors: Hui Chen, Lirong YangAbstract:The resolutions of racemic diastereomeric mixtures of menthyl propionate was performed by Pseudomonas Alcaligenes lipase (PaL) to produce (2S, 5R) L-menthol. Because of the inherently low diastereopreference of PaL, covalent docking and molecular dynamic (MD) simulations were used to investigate possible avenues of improvement. Rational site-directed mutagenesis of PaL revealed residues V180 and A272 to be the hotspots for diastereopreference. The double V180L/A272F mutant exhibited the highest degree of diastereopreference, as the diastereomeric ratio of (2S, 5R) L-menthol increased towards both (2R, 5S) L-neomenthol (dr1) and (2R, 5R) D-isoneomenthol (dr2) (diastereomeric ratios dr1 and dr2 increased to 4.65 and 2.13 times that of wild-type PaL). MD simulation analysis indicated that these mutations decrease the flexibility of the surrounding protein regions. The combination of increased steric exclusion and decreased flexibility results in less favorable binding of the non-target substrates, (2R, 5S) L-neomenthyl propionate and (2R, 5R) D-isoneomenthyl propionate, to the V180L/A272F mutant. These results confirmed and further improved our previously proposed model of the diastereomer recognition mechanism based on the combined effect of steric exclusion and regional flexibility.
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characterization and structure basis of Pseudomonas Alcaligenes lipase s enantiopreference towards d l menthyl propionate
Journal of Molecular Catalysis B-enzymatic, 2014Co-Authors: Hui Chen, Lirong YangAbstract:Abstract In this >work, a lipase from Pseudomonas Alcaligenes CGMCC4405 (PaL) was cloned and expressed. It was very attractive that the recombinant PaL exhibited excellent enantioselectivity ( E > 200) in the resolution of racemic d , l -menthyl propionate to produce l -menthol. The structure basis of enantiopreference is a fundamental scientific problem which needs to be resolved. In our research, molecular dynamic simulation (MD) research was employed to research the different binding modes of d and l -menthyl propionate. The results showed that when bound with slow-reacting enantiomer ( d -menthyl propionate), the steric requirements of the large substituent (isopropyl) of the d -menthyl propionate force a rotation of the imidazole ring of catalytic residue His271 and further pushed the active site His271 away from its proper orientation. Moreover, the average distance between alcohol oxygen (O alc ) and H Nɛ of catalytic His271 increased to 3.7 A, which was too far to form an essential hydrogen bond and further prevented efficient catalysis of slow enantiomer. This correlation of the distance between alcohol oxygen (O alc ) and H Nɛ of catalytic His271 and the enantioselectivity was also confirmed by the result of site-directed mutagenesis.
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A combination of site-directed mutagenesis and chemical modification to improve diastereopreference of Pseudomonas Alcaligenes lipase.
Biochimica et biophysica acta, 2013Co-Authors: Hui Chen, Lirong YangAbstract:Abstract A combination of site-directed mutagenesis and chemical modification was employed to alter protein structure with the objective of improving diastereopreference over that achieved by simple site-directed mutagenesis. Conformational analysis using molecular dynamic (MD) simulation of Pseudomonas Alcaligenes lipase (PAL) indicated that stronger steric exclusion and structural rigidity facilitated diastereopreference. A cysteine (Cys) residue was introduced using site-directed mutagenesis to construct variant A272C. The modifier 5,5′-dithiobis-(2-nitrobenzoic acid) (DTNB) was then reacted with the introduced Cys residue to provide stronger steric exclusion and structural rigidity. The modification was verified by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Diastereopreference was improved significantly. The diastereomeric excess (dep) of l -menthol increased from 35% with wild type PAL to 90% with A272C-DTNB modified PAL when the conversion ratio of l -menthyl propionate was nearly 100%. Conformation and kinetic parameter analysis showed that A272C-DTNB modified PAL exhibited stronger steric exclusion and increased structural rigidity around the modification site that inhibited the hydrolysis of non-targeted substrates. The combination of site-directed mutagenesis and chemical modification could be an effective method to alter protein properties and enhance diastereopreference through the combined effect of steric exclusion and structural rigidity.
Wim J Quax - One of the best experts on this subject based on the ideXlab platform.
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lipase a gene transcription in Pseudomonas Alcaligenes is under control of rna polymerase σ54 and response regulator lipr
Fems Microbiology Letters, 2012Co-Authors: Joanna Krzeslak, Ronald Van Merkerk, Robbert H Cool, Evelina Papaioannou, Krisztina Paal, Rainer Bischoff, Wim J QuaxAbstract:Initial analysis has shown that the transcription of the Pseudomonas Alcaligenes lipA gene, which encodes an extracellular lipase, is governed by the LipQR two-component system consisting of sensor kinase LipQ and DNA-binding regulator LipR. This study further analyzes lipA gene expression and demonstrates that the RNA polymerase σ54 is involved in the transcription. Purified LipR has an ATPase activity that is stimulated by the presence of lipA promoter DNA. Surface plasmon resonance measurements with purified and in vitro phosphorylated LipR reveal that phosphorylation of LipR is required for specific binding to the upstream activating sequence of the lipA promoter. Furthermore, mass spectrometric analysis combined with mutagenesis demonstrates that Asp52 is the phosphorylated aspartate. This analysis exposes LipR as a prominent member of the growing family of bacterial enhancer-binding proteins.
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lipase expression in Pseudomonas Alcaligenes is under the control of a two component regulatory system
Applied and Environmental Microbiology, 2008Co-Authors: Joanna Krzeslak, Gijs Gerritse, Wim J Quax, Ronald Van Merkerk, Robbert H CoolAbstract:Preliminary observations in a large-scale fermentation process suggested that the lipase expression of Pseudomonas Alcaligenes can be switched on by the addition of certain medium components, such as soybean oil. In an attempt to elucidate the mechanism of induction of lipase expression, we have set up a search method for genes controlling lipase expression by use of a cosmid library containing fragments of P. Alcaligenes genomic DNA. A screen for lipase hyperproduction resulted in the selection of multiple transformants, of which the best-producing strains comprised cosmids that shared an overlapping genomic fragment. Within this fragment, two previously unidentified genes were found and named lipQ and lipR. Their encoded proteins belong to the NtrBC family of regulators that regulate gene expression via binding to a specific upstream activator sequence (UAS). Such an NtrC-like UAS was identified in a previous study in the P. Alcaligenes lipase promoter, strongly suggesting that LipR acts as a positive regulator of lipase expression. The regulating role could be confirmed by down-regulated lipase expression in a strain with an inactivated lipR gene and a threefold increase in lipase yield in a large-scale fermentation when expressing the lipQR operon from the multicopy plasmid pLAFR3. Finally, cell extracts of a LipR-overexpressing strain caused a retardation of the lipase promoter fragment in a band shift assay. Our results indicate that lipase expression in Pseudomonas Alcaligenes is under the control of the LipQR two-component system.
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Characterization of the promoter and upstream activating sequence from the Pseudomonas Alcaligenes lipase gene.
Journal of biotechnology, 2001Co-Authors: Manon M.j. Cox, Gijs Gerritse, Lydia Dankmeyer, Wim J QuaxAbstract:Pseudomonas Alcaligenes secretes a lipase with a high pH optimum, which has interesting properties for application in detergents. The expression of the lipase is strongly dependent on the presence of lipids in the growth medium such as soybean oil. The promoter of the gene was characterized and found to have resemblance to sigma 54 controlled promoters, which are known to be tightly regulated. The transcription start was mapped precisely downstream of a sequence with close similarity to the -12/-24 consensus sequence of sigma 54 controlled promoters. Interestingly, a hyperproducer mutant strain was isolated and found to have a C to T mutation in the -12/-24 promoter consensus region. In addition an Upstream Activating Sequence (UAS) with homology to sigma 54 UAS consensus sequences was identified. It was demonstrated that an increase of the distance from the UAS to the transcription start or the deletion of the UAS results in significantly lower expression levels of lipase. A systematic mutational analysis of the UAS sequence has resulted in a variant with an increased lipase expression. (C) 2001 Elsevier Science B.V. All rights reserved.
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the phenotype enhancement method identifies the xcp outer membrane secretion machinery from Pseudomonas Alcaligenes as a bottleneck for lipase production
Journal of Biotechnology, 1998Co-Authors: Gijs Gerritse, Roisin Ure, Fanny Bizoullier, Wim J QuaxAbstract:Pseudomonas Alcaligenes M-1 has been selected from an intensive screening for micro-organisms that can naturally produce a lipase active in detergent formulations. The lipase expression has been increased to allow high level secretion from Pseudomonas Alcaligenes, via the introduction of multi-copy plasmids. In order to improve the lipase yield further, the phenotype enhancement method has been developed. This idea comprises the reintroduction of a cosmid library with random chromosomal fragments in a P. Alcaligenes strain with already high lipase productivity. One of the strains which showed an enhanced lipase production appeared to contain a cosmid encoding the outer membrane secretion genes. These xcp-genes are clustered in two divergently transcribed operons similar to the situation in Pseudomonas aeruginosa. Remarkably and dissimilar to P. aeruginosa, in between the two xcp gene clusters, two reading frames of unknown function--OrfV and OrfX--are present. For OrfX no equivalent can be found in the known protein data bases. On the other hand, OrfV shows homology to the regulatory proteins MalT and AcoK. Some evidence is provided that suggests that OrfV acts as a regulator of the xcp operons. A model is proposed for the regulation of the secretion system from P. Alcaligenes.
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Development of a Lipase Fermentation Process That Uses a Recombinant Pseudomonas Alcaligenes Strain
Applied and Environmental Microbiology, 1998Co-Authors: Gijs Gerritse, Ronald Hommes, Wim J QuaxAbstract:Pseudomonas Alcaligenes M-l secretes an alkaline lipase, which has excellent characteristics for the removal of fatty stains under modern washing conditions. A fed-batch fermentation process based on the secretion of the alkaline lipase from P. Alcaligenes was developed. Due to the inability of P. Alcaligenes to grow on glucose, citric acid and soybean oil were applied as substrates in the batch phase and feed phase, respectively. The gene encoding the high-alkaline lipase from P. Alcaligenes was isolated and characterized. Amplification of lipase gene copies in P. Alcaligenes with the aid of low- and high-copy-number plasmids resulted in an increase of lipase expression that was apparently colinear,vith the gene copy number, It was found that overexpression of the lipase helper gene, lipB, produced a stimulating effect in strains with high copy numbers (>20) of the lipase structural gene, lipA. In strains with lipA on a low-copy-number vector, the lipB gene did not show any effect, suggesting that LipB is required in a low ratio to LipA only, During scaling up of the fermentation process to 100 m(3), severe losses in lipase productivity were observed. Simulations have identified an increased level of dissolved carbon dioxide as the most probable cause for the scale-up losses. A large-scale fermentation protocol with a reduced dissolved carbon dioxide concentration resulted in a substantial elimination of the scale-up loss.
Zaki A Siddiqui - One of the best experts on this subject based on the ideXlab platform.
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biocontrol of fusarium wilt by bacillus pumilus Pseudomonas Alcaligenes and rhizobium sp on lentil
Turkish Journal of Biology, 2010Co-Authors: Mohd Sayeed Akhtar, Uzma Shakeel, Zaki A SiddiquiAbstract:The present study examined the effects of Bacillus pumilus, Pseudomonas Alcaligenes, and Rhizobium sp. on wilt disease caused by Fusarium oxysporum f. sp. lentis and on the growth of lentil. Inoculation with F. oxysporum caused significant wilting, and reduced plant growth, the number of pods, and nodulation. Inoculation with B. pumilus together with P. Alcaligenes caused a greater increase in plant growth, number of pods, nodulation, and root colonization by rhizobacteria, and also reduced Fusarium wilting to a greater degree than did individual inoculation. Use of Rhizobium sp. resulted in a greater increase in plant growth, number of pods, and nodulation, and reduced wilting more than B. pumilus did. Combined application of B. pumilus and P. Alcaligenes with Rhizobium sp. resulted in the greatest increase in plant growth, number of pods, nodulation, and root colonization by rhizobacteria, and also reduced wilting in Fusarium-inoculated plants.
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use of plant growth promoting rhizobacteria for the biocontrol of root rot disease complex of chickpea
Australasian Plant Pathology, 2009Co-Authors: Mohd Sayeed Akhtar, Zaki A SiddiquiAbstract:The effects of Pseudomonas putida, Pseudomonas Alcaligenes and a Pseudomonas isolate (Ps28) on the hatching and penetration of Meloidogyne incognita in chickpea (Cicer arietinum) roots were studied. Root colonisation, antifungal activity against Macrophomina phaseolina and the production of siderophores, hydrogen cyanide (HCN) and indole acetic acid (IAA) were also estimated for each bacterial isolate. P. putida had the greatest inhibitory effect on hatching and root penetration of M. incognita followed by P. Alcaligenes and Ps28, respectively. Similarly, P. putida colonised roots more effectively than P. Alcaligenes or Ps28. In addition, P. putida had the greatest inhibitory effect on M. Phaseolina and produced the greatest amounts of siderophores, IAA and HCN compared with P. Alcaligenes and Ps28. The effects of these bacterial isolates on plant growth and root-rot disease complex of chickpea caused by M. Incognita and M. phaseolina were observed. Plant inoculations with these bacterial isolates increased plant growth and the number of seed pods in diseased plants while reducing galling, nematode multiplication and the root-rot disease index. P. putida caused the greatest reduction in galling and nematode multiplication followed by P. Alcaligenes and Ps28, respectively. The present study suggests that P. putida has potential for the biocontrol of root-rot disease complex of chickpea.
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glomus intraradices Pseudomonas Alcaligenes and bacillus pumilus effective agents for the control of root rot disease complex of chickpea cicer arietinum l
Journal of General Plant Pathology, 2008Co-Authors: Sayeed M Akhtar, Zaki A SiddiquiAbstract:The effects of Glomus intraradices, Pseudomonas Alcaligenes and Bacillus pumilus on the root-rot disease complex caused by the root-knot nematode Meloidogyne incognita and the root-rot fungus Macrophomina phaseolina in chickpea was assessed by quantifying differences in the shoot dry mass, pod number, nodulation, and shoot content of chlorophyll, nitrogen, phosphorus and potassium. Inoculation of plants with G. intraradices, P. Alcaligenes and B. pumilus alone and in combination significantly increased shoot dry mass, pod number, and content of chlorophyll, nitrogen, phosphorus and potassium in plants inoculated with pathogens over that in the uninoculated control plants. P. Alcaligenes caused a greater increase in shoot dry mass, pod number, chlorophyll, nitrogen, phosphorus and potassium in plants with pathogens than did G. intraradices or B. pumilus. Combined application of G. intraradices, P. Alcaligenes and B. pumilus to plants inoculated with pathogens caused a greater increase in shoot dry mass, pod number, nitrogen, phosphorus, and potassium than did an application of P. Alcaligenes plus B. pumilus or of G. intraradices plus B. pumilus. Root colonization by G. intrardices was high when used alone, while inoculation with the pathogens reduced root colonization by G. intraradices. In the presence of P. Alcaligenes and B. pumilus, root colonization by G. intraradices increased. In plants inoculated with just one antagonist, P. Alcaligenes reduced galling and nematode multiplication the most, followed by G. intraradices, then B. pumilus. The greatest reduction in galling, nematode multiplication and root-rot was observed when both bacterial species and G. intraradices were applied together.
