The Experts below are selected from a list of 3003 Experts worldwide ranked by ideXlab platform
Zhen-ming Chi - One of the best experts on this subject based on the ideXlab platform.
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Inulinase production by the yeast Kluyveromyces marxianus with the disrupted MIG1 gene and the over-expressed Inulinase gene
Process Biochemistry, 2014Co-Authors: Hai-xiang Zhou, Guang-lei Liu, Zhe Chi, Fang-hua Xin, Zhen-ming ChiAbstract:Kluyveromyces marxianus has been widely used in food industries and Inulinase produced by the yeast has many applications. In order to greatly enhance Inulinase production by the yeast, it is very important to both derepress glucose repression and overexpress the native Inulinase gene. The MIG1 gene in the yeast K. marxianus KM-0 was disrupted. It was found that a glucose-derepressed mutant (KM-69) of K. marxianus KM-0 could produce 94.6 units of Inulinase activity per ml in the inulin medium within 36 h. In order to further increase Inulinase production, the native Inulinase gene was over-expressed in the glucose-derepressed mutant. During the 10-l fermentation, the recombinant strain KM-526 yielded 133.5 units of Inulinase activity per ml within very short time (24 h). The results also showed that Inulinase produced by the recombinant strain KM-526 could more effectively convert inulin into monosaccharides than that produced by K. marxianus KM-0. The results indicate that it is very important to delete the MIG1 gene in K. marxianus KM-0 and over-express the native INU1 gene in order to further enhance Inulinase yield within the short time by this yeast and for efficient hydrolysis of inulin. The recombinant strain KM-526 and its Inulinase have highly potential applications in food industries.
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Molecular characterization and expression of microbial Inulinase genes
Critical reviews in microbiology, 2012Co-Authors: Guang-lei Liu, Zhe Chi, Zhen-ming ChiAbstract:Many genes encoding exo- and endo-Inulinases from bacteria, yeasts and filamentous fungi have been cloned and characterized. All the Inulinases have several conserved motifs, such as WMND(E)PNGL, RDP, EC(V)P, SVEVF, Q and FS(T), which play an important role in Inulinase catalysis and substrate binding. However, the exo-Inulinases produced by yeasts has no conserved motif SVEVF and the yeasts do not produce any endo-Inulinase. Exo- and endo-Inulinases found in different microorganisms cluster separately at distant positions from each other. Most of the cloned Inulinase genes have been expressed in Yarrowia lipolytica, Saccharomyces cerevisiae, Pichia pastoris, Klyuveromyces lactis and Escherichia coli, respectively. The recombinant Inulinases produced and the engineered hosts using the cloned Inulinase genes have many potential applications. Expression of most of the Inulinase genes is repressed by glucose and fructose and induced by inulin and sucrose. However, the detailed mechanisms of the repression an...
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enhanced Inulinase production in solid state fermentation by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Journal of Industrial Microbiology & Biotechnology, 2009Co-Authors: Ning Guo, Zhen-ming Chi, Fang Gong, Jun ShengAbstract:In order to isolate Inulinase overproducers of the marine yeast Pichia guilliermondii, strain 1, cells were mutated by using UV light and LiCl2. One mutant (M-30) with enhanced Inulinase production was obtained. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant in solid-state fermentation. The initial moisture, inoculum, the amount ratio of wheat bran to rice bran, temperature, pH for the maximum Inulinase production by the mutant M-30 were found to be 60.5%, 2.5%, 0.42, 30°C and 6.50, respectively. Under the optimized conditions, 455.9 U/grams of dry substrate (gds) of Inulinase activity was reached in the solid state fermentation culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 459.2 U/gds was derived from RSM regression. Under the same conditions, its parent strain only produced 291.0 U/gds of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far.
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use of response surface methodology for optimizing process parameters for high Inulinase production by the marine yeast cryptococcus aureus g7a in solid state fermentation and hydrolysis of inulin
Bioprocess and Biosystems Engineering, 2009Co-Authors: Jun Sheng, Zhen-ming Chi, Xianghong Wang, Kuirang Yan, Fang GongAbstract:The optimization of process parameters for high Inulinase production by the marine yeast strain Cryptococcus aureus G7a in solid-state fermentation (SSF) was carried out using central composite design (CCD), one of the response surface methodologies (RSMs). We found that moisture, inoculation size, the amount ratio of wheat bran to rice husk, temperature and pH had great influence on Inulinase production by strain G7a. Therefore, the CCD was used to evaluate the influence of the five factors on the Inulinase production by strain G7a. Then, five levels of the five factors above were further optimized using the CCD. Finally, the optimal parameters obtained with the RSM were the initial moisture 61.5%, inoculum 2.75%, the amount ratio of wheat bran to rice husk 0.42, temperature 29 °C, pH 5.5. Under the optimized conditions, 420.9 U g−1 of dry substrate of Inulinase activity was reached in the solid-state fermentation culture of strain G7a within 120 h whereas the predicted maximum Inulinase activity of 436.2 U g−1 of Inulinase activity of 436.2 U g−1 of dry weight was derived from the RSM regression. This is the highest Inulinase activity produced by the yeast strain reported so far. A large amount of monosaccharides and oligosaccharides were detected after inulin hydrolysis by the crude Inulinase.
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Inulinase overproduction by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Biochemical Engineering Journal, 2009Co-Authors: Ning Guo, Fang Gong, Zhen-ming Chi, Jun Sheng, Zhe ChiAbstract:Abstract In this study, in order to isolate Inulinase overproducers from the marine yeast Pichia guilliermondii, its cells were treated by using UV light and LiCl. The mutant M-30 with enhanced Inulinase production was obtained and was found to be stable after cultivation for 20 generations. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant M-30 in liquid fermentation. Inulin, yeast extract, NaCl, temperature, pH for maximum Inulinase production by the mutant M-30 were found to be 20.0 g/l, 5.0 g/l, 20.0 g/l, 28 °C and 6.5, respectively. Under the optimized conditions, 127.7 U/ml of Inulinase activity was reached in the liquid culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 129.8 U/ml was derived from RSM regression. Under the same conditions, its parent strain only produced 48.1 U/ml of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far. We also found that inulin could be actively converted into monosaccharides by the crude Inulinase.
Fang Gong - One of the best experts on this subject based on the ideXlab platform.
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use of response surface methodology for optimizing process parameters for high Inulinase production by the marine yeast cryptococcus aureus g7a in solid state fermentation and hydrolysis of inulin
Bioprocess and Biosystems Engineering, 2009Co-Authors: Jun Sheng, Zhen-ming Chi, Xianghong Wang, Kuirang Yan, Fang GongAbstract:The optimization of process parameters for high Inulinase production by the marine yeast strain Cryptococcus aureus G7a in solid-state fermentation (SSF) was carried out using central composite design (CCD), one of the response surface methodologies (RSMs). We found that moisture, inoculation size, the amount ratio of wheat bran to rice husk, temperature and pH had great influence on Inulinase production by strain G7a. Therefore, the CCD was used to evaluate the influence of the five factors on the Inulinase production by strain G7a. Then, five levels of the five factors above were further optimized using the CCD. Finally, the optimal parameters obtained with the RSM were the initial moisture 61.5%, inoculum 2.75%, the amount ratio of wheat bran to rice husk 0.42, temperature 29 °C, pH 5.5. Under the optimized conditions, 420.9 U g−1 of dry substrate of Inulinase activity was reached in the solid-state fermentation culture of strain G7a within 120 h whereas the predicted maximum Inulinase activity of 436.2 U g−1 of Inulinase activity of 436.2 U g−1 of dry weight was derived from the RSM regression. This is the highest Inulinase activity produced by the yeast strain reported so far. A large amount of monosaccharides and oligosaccharides were detected after inulin hydrolysis by the crude Inulinase.
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enhanced Inulinase production in solid state fermentation by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Journal of Industrial Microbiology & Biotechnology, 2009Co-Authors: Ning Guo, Zhen-ming Chi, Fang Gong, Jun ShengAbstract:In order to isolate Inulinase overproducers of the marine yeast Pichia guilliermondii, strain 1, cells were mutated by using UV light and LiCl2. One mutant (M-30) with enhanced Inulinase production was obtained. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant in solid-state fermentation. The initial moisture, inoculum, the amount ratio of wheat bran to rice bran, temperature, pH for the maximum Inulinase production by the mutant M-30 were found to be 60.5%, 2.5%, 0.42, 30°C and 6.50, respectively. Under the optimized conditions, 455.9 U/grams of dry substrate (gds) of Inulinase activity was reached in the solid state fermentation culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 459.2 U/gds was derived from RSM regression. Under the same conditions, its parent strain only produced 291.0 U/gds of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far.
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Inulinase overproduction by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Biochemical Engineering Journal, 2009Co-Authors: Ning Guo, Fang Gong, Zhen-ming Chi, Jun Sheng, Zhe ChiAbstract:Abstract In this study, in order to isolate Inulinase overproducers from the marine yeast Pichia guilliermondii, its cells were treated by using UV light and LiCl. The mutant M-30 with enhanced Inulinase production was obtained and was found to be stable after cultivation for 20 generations. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant M-30 in liquid fermentation. Inulin, yeast extract, NaCl, temperature, pH for maximum Inulinase production by the mutant M-30 were found to be 20.0 g/l, 5.0 g/l, 20.0 g/l, 28 °C and 6.5, respectively. Under the optimized conditions, 127.7 U/ml of Inulinase activity was reached in the liquid culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 129.8 U/ml was derived from RSM regression. Under the same conditions, its parent strain only produced 48.1 U/ml of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far. We also found that inulin could be actively converted into monosaccharides by the crude Inulinase.
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Optimization for high-level expression of the Pichia guilliermondii recombinant Inulinase in Pichia pastoris and characterization of the recombinant Inulinase
Process Biochemistry, 2009Co-Authors: Tong Zhang, Fang Gong, Ying Peng, Zhen-ming ChiAbstract:Abstract The Inulinase gene cloned from the marine-derived yeast Pichia guilliermondii strain 1 was expressed in Pichia pastoris X-33 and the conditions for overexpression of the Inulinase were optimized. After the optimization of the conditions for production of the recombinant Inulinase, 286.8 ± 5.4 U/ml and 8873 ± 55.3 U/mg of the recombinanat Inulinase in the supernatant of the culture of 2-l fermentor were attained at 120 h of the fermentation and fermentation efficiency was 13.04 μg ± 0.4 of protein/ml/d. The recombinant Inulinase was purified and characterized. The molecular weight of the purified recombinant Inulinase was 57.6 kDa, which was higher than that of the native iunlinase. The optimal pH and temperature of the purified recombinant Inulinase were 6.0 and 60 °C, respectively. Other biochemical characteristics of the purified recombinant Inulinase were the same as those of the native Inulinase produced by the marine-derived P. guilliermondii strain 1. The purified recombinant Inulinase also had high exoInulinase activity. Therefore, the recombinant Inulinase may have highly potential applications in food and pharmaceutical industies.
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cloning and characterization of the Inulinase gene from a marine yeast pichia guilliermondii and its expression in pichia pastoris
Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology, 2009Co-Authors: Tong Zhang, Guang-lei Liu, Zhe Chi, Zhen-ming Chi, Fang Gong, Jun Sheng, Xianghong WangAbstract:The extracellular Inulinase structural gene was isolated from the genomic DNA of the marine yeast Pichia guilliermondii strain 1 by PCR. The gene had an open reading frame of 1,542 bp long encoding an Inulinase. The coding region of the gene was not interrupted by any intron. It encoded 514 amino acid residues of a protein with a putative signal peptide of 18 amino acids and the calculated molecular mass of 58.04 kDa. The protein sequence deduced from the Inulinase structural gene contained the Inulinase consensus sequences (WMNXPNGL) and (RDPKVF). It also had ten conserved putative N-glycosylation sites. The Inulinase from P. guilliermondii strain 1 was found to be closely related to that from Kluyveromyces marxianus. The Inulinase gene without the signal sequence was subcloned into pPICZαA expression vector and expressed in Pichia pastoris X-33. The expressed fusion protein was analyzed by SDS-PAGE and western blotting and a specific band with molecular mass of about 60 kDa was found. Enzyme activity assay verified the recombinant protein as an Inulinase. A maximum activity of 58.7 ± 0.12 U/ml was obtained from the culture supernatant of P. pastoris X-33 harboring the Inulinase gene. A large amount of monosaccharides, disaccharides and oligosaccharides were detected after the hydrolysis of inulin with the crude recombinant Inulinase.
Jun Sheng - One of the best experts on this subject based on the ideXlab platform.
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enhanced Inulinase production in solid state fermentation by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Journal of Industrial Microbiology & Biotechnology, 2009Co-Authors: Ning Guo, Zhen-ming Chi, Fang Gong, Jun ShengAbstract:In order to isolate Inulinase overproducers of the marine yeast Pichia guilliermondii, strain 1, cells were mutated by using UV light and LiCl2. One mutant (M-30) with enhanced Inulinase production was obtained. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant in solid-state fermentation. The initial moisture, inoculum, the amount ratio of wheat bran to rice bran, temperature, pH for the maximum Inulinase production by the mutant M-30 were found to be 60.5%, 2.5%, 0.42, 30°C and 6.50, respectively. Under the optimized conditions, 455.9 U/grams of dry substrate (gds) of Inulinase activity was reached in the solid state fermentation culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 459.2 U/gds was derived from RSM regression. Under the same conditions, its parent strain only produced 291.0 U/gds of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far.
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use of response surface methodology for optimizing process parameters for high Inulinase production by the marine yeast cryptococcus aureus g7a in solid state fermentation and hydrolysis of inulin
Bioprocess and Biosystems Engineering, 2009Co-Authors: Jun Sheng, Zhen-ming Chi, Xianghong Wang, Kuirang Yan, Fang GongAbstract:The optimization of process parameters for high Inulinase production by the marine yeast strain Cryptococcus aureus G7a in solid-state fermentation (SSF) was carried out using central composite design (CCD), one of the response surface methodologies (RSMs). We found that moisture, inoculation size, the amount ratio of wheat bran to rice husk, temperature and pH had great influence on Inulinase production by strain G7a. Therefore, the CCD was used to evaluate the influence of the five factors on the Inulinase production by strain G7a. Then, five levels of the five factors above were further optimized using the CCD. Finally, the optimal parameters obtained with the RSM were the initial moisture 61.5%, inoculum 2.75%, the amount ratio of wheat bran to rice husk 0.42, temperature 29 °C, pH 5.5. Under the optimized conditions, 420.9 U g−1 of dry substrate of Inulinase activity was reached in the solid-state fermentation culture of strain G7a within 120 h whereas the predicted maximum Inulinase activity of 436.2 U g−1 of Inulinase activity of 436.2 U g−1 of dry weight was derived from the RSM regression. This is the highest Inulinase activity produced by the yeast strain reported so far. A large amount of monosaccharides and oligosaccharides were detected after inulin hydrolysis by the crude Inulinase.
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Inulinase overproduction by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Biochemical Engineering Journal, 2009Co-Authors: Ning Guo, Fang Gong, Zhen-ming Chi, Jun Sheng, Zhe ChiAbstract:Abstract In this study, in order to isolate Inulinase overproducers from the marine yeast Pichia guilliermondii, its cells were treated by using UV light and LiCl. The mutant M-30 with enhanced Inulinase production was obtained and was found to be stable after cultivation for 20 generations. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant M-30 in liquid fermentation. Inulin, yeast extract, NaCl, temperature, pH for maximum Inulinase production by the mutant M-30 were found to be 20.0 g/l, 5.0 g/l, 20.0 g/l, 28 °C and 6.5, respectively. Under the optimized conditions, 127.7 U/ml of Inulinase activity was reached in the liquid culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 129.8 U/ml was derived from RSM regression. Under the same conditions, its parent strain only produced 48.1 U/ml of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far. We also found that inulin could be actively converted into monosaccharides by the crude Inulinase.
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cloning and characterization of the Inulinase gene from a marine yeast pichia guilliermondii and its expression in pichia pastoris
Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology, 2009Co-Authors: Tong Zhang, Guang-lei Liu, Zhe Chi, Zhen-ming Chi, Fang Gong, Jun Sheng, Xianghong WangAbstract:The extracellular Inulinase structural gene was isolated from the genomic DNA of the marine yeast Pichia guilliermondii strain 1 by PCR. The gene had an open reading frame of 1,542 bp long encoding an Inulinase. The coding region of the gene was not interrupted by any intron. It encoded 514 amino acid residues of a protein with a putative signal peptide of 18 amino acids and the calculated molecular mass of 58.04 kDa. The protein sequence deduced from the Inulinase structural gene contained the Inulinase consensus sequences (WMNXPNGL) and (RDPKVF). It also had ten conserved putative N-glycosylation sites. The Inulinase from P. guilliermondii strain 1 was found to be closely related to that from Kluyveromyces marxianus. The Inulinase gene without the signal sequence was subcloned into pPICZαA expression vector and expressed in Pichia pastoris X-33. The expressed fusion protein was analyzed by SDS-PAGE and western blotting and a specific band with molecular mass of about 60 kDa was found. Enzyme activity assay verified the recombinant protein as an Inulinase. A maximum activity of 58.7 ± 0.12 U/ml was obtained from the culture supernatant of P. pastoris X-33 harboring the Inulinase gene. A large amount of monosaccharides, disaccharides and oligosaccharides were detected after the hydrolysis of inulin with the crude recombinant Inulinase.
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Purification and characterization of extracellular Inulinase from a marine yeast Pichia guilliermondii and inulin hydrolysis by the purified Inulinase
Biotechnology and Bioprocess Engineering, 2008Co-Authors: Fang Gong, Zhen-ming Chi, Tong Zhang, Jun Sheng, Xianghong WangAbstract:The extracellular Inulinase of the marine yeast Pichia guilliermondii strain 1 was purified to homogeneity resulting in a 7.2-fold increase in specific Inulinase activity. The molecular mass of the purified enzyme was estimated to be 50.0 kDa. The optimal pH and temperature for the purified enzyme were 6.0 and 60°C, respectively. The enzyme was activated by Mn2+, Ca2+, K+, Li+, Na+, Fe3+, Fe2+, Cu2+, and Co2+, but Mg2+, Hg2+, and Ag+ inhibited activity. The enzyme was strongly inhibited by phenylmethanesulphonyl fluoride (PMSF), iodoacetic acid, EDTA, and 1, 10-phenanthroline. The Km and Vmax values of the purified Inulinase for inulin were 21.1 mg/mL and 0.08 mg/min, respectively. A large number of monosaccharides were detected after the hydrolysis of inulin. The deduced protein sequence from the cloned P. guilliermondii strain 1 Inulinase gene contained the consensus motifs R-D-P-K-V-F-W-H and W-M-N-D-P-N-G, which are conserved among the Inulinases from other microorganisms.
Zhe Chi - One of the best experts on this subject based on the ideXlab platform.
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Inulinase production by the yeast Kluyveromyces marxianus with the disrupted MIG1 gene and the over-expressed Inulinase gene
Process Biochemistry, 2014Co-Authors: Hai-xiang Zhou, Guang-lei Liu, Zhe Chi, Fang-hua Xin, Zhen-ming ChiAbstract:Kluyveromyces marxianus has been widely used in food industries and Inulinase produced by the yeast has many applications. In order to greatly enhance Inulinase production by the yeast, it is very important to both derepress glucose repression and overexpress the native Inulinase gene. The MIG1 gene in the yeast K. marxianus KM-0 was disrupted. It was found that a glucose-derepressed mutant (KM-69) of K. marxianus KM-0 could produce 94.6 units of Inulinase activity per ml in the inulin medium within 36 h. In order to further increase Inulinase production, the native Inulinase gene was over-expressed in the glucose-derepressed mutant. During the 10-l fermentation, the recombinant strain KM-526 yielded 133.5 units of Inulinase activity per ml within very short time (24 h). The results also showed that Inulinase produced by the recombinant strain KM-526 could more effectively convert inulin into monosaccharides than that produced by K. marxianus KM-0. The results indicate that it is very important to delete the MIG1 gene in K. marxianus KM-0 and over-express the native INU1 gene in order to further enhance Inulinase yield within the short time by this yeast and for efficient hydrolysis of inulin. The recombinant strain KM-526 and its Inulinase have highly potential applications in food industries.
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Molecular characterization and expression of microbial Inulinase genes
Critical reviews in microbiology, 2012Co-Authors: Guang-lei Liu, Zhe Chi, Zhen-ming ChiAbstract:Many genes encoding exo- and endo-Inulinases from bacteria, yeasts and filamentous fungi have been cloned and characterized. All the Inulinases have several conserved motifs, such as WMND(E)PNGL, RDP, EC(V)P, SVEVF, Q and FS(T), which play an important role in Inulinase catalysis and substrate binding. However, the exo-Inulinases produced by yeasts has no conserved motif SVEVF and the yeasts do not produce any endo-Inulinase. Exo- and endo-Inulinases found in different microorganisms cluster separately at distant positions from each other. Most of the cloned Inulinase genes have been expressed in Yarrowia lipolytica, Saccharomyces cerevisiae, Pichia pastoris, Klyuveromyces lactis and Escherichia coli, respectively. The recombinant Inulinases produced and the engineered hosts using the cloned Inulinase genes have many potential applications. Expression of most of the Inulinase genes is repressed by glucose and fructose and induced by inulin and sucrose. However, the detailed mechanisms of the repression an...
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Inulinase overproduction by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Biochemical Engineering Journal, 2009Co-Authors: Ning Guo, Fang Gong, Zhen-ming Chi, Jun Sheng, Zhe ChiAbstract:Abstract In this study, in order to isolate Inulinase overproducers from the marine yeast Pichia guilliermondii, its cells were treated by using UV light and LiCl. The mutant M-30 with enhanced Inulinase production was obtained and was found to be stable after cultivation for 20 generations. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant M-30 in liquid fermentation. Inulin, yeast extract, NaCl, temperature, pH for maximum Inulinase production by the mutant M-30 were found to be 20.0 g/l, 5.0 g/l, 20.0 g/l, 28 °C and 6.5, respectively. Under the optimized conditions, 127.7 U/ml of Inulinase activity was reached in the liquid culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 129.8 U/ml was derived from RSM regression. Under the same conditions, its parent strain only produced 48.1 U/ml of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far. We also found that inulin could be actively converted into monosaccharides by the crude Inulinase.
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Inulinase-expressing microorganisms and applications of Inulinases.
Applied microbiology and biotechnology, 2009Co-Authors: Zhen-ming Chi, Guang-lei Liu, Zhe Chi, Tong Zhang, Lixi YueAbstract:In this review article, Inulinase-expressing microorganisms and its potential applications in transformation of inulin into very-high-fructose syrup, bioethanol, and inulooligosaccharides are overviewed. In the past 10 years, many new Inulinase producers have been obtained and many genes encoding Inulinases from different microorganisms have been cloned and characterized. Some novel processes for exoInulinase overproduction have been developed for bioethanol production and ultra-high-fructose syrup. The endoInulinases have also been used for production of inulooligosaccharides from inulin and inulin-containing materials.
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cloning and characterization of the Inulinase gene from a marine yeast pichia guilliermondii and its expression in pichia pastoris
Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology, 2009Co-Authors: Tong Zhang, Guang-lei Liu, Zhe Chi, Zhen-ming Chi, Fang Gong, Jun Sheng, Xianghong WangAbstract:The extracellular Inulinase structural gene was isolated from the genomic DNA of the marine yeast Pichia guilliermondii strain 1 by PCR. The gene had an open reading frame of 1,542 bp long encoding an Inulinase. The coding region of the gene was not interrupted by any intron. It encoded 514 amino acid residues of a protein with a putative signal peptide of 18 amino acids and the calculated molecular mass of 58.04 kDa. The protein sequence deduced from the Inulinase structural gene contained the Inulinase consensus sequences (WMNXPNGL) and (RDPKVF). It also had ten conserved putative N-glycosylation sites. The Inulinase from P. guilliermondii strain 1 was found to be closely related to that from Kluyveromyces marxianus. The Inulinase gene without the signal sequence was subcloned into pPICZαA expression vector and expressed in Pichia pastoris X-33. The expressed fusion protein was analyzed by SDS-PAGE and western blotting and a specific band with molecular mass of about 60 kDa was found. Enzyme activity assay verified the recombinant protein as an Inulinase. A maximum activity of 58.7 ± 0.12 U/ml was obtained from the culture supernatant of P. pastoris X-33 harboring the Inulinase gene. A large amount of monosaccharides, disaccharides and oligosaccharides were detected after the hydrolysis of inulin with the crude recombinant Inulinase.
Ning Guo - One of the best experts on this subject based on the ideXlab platform.
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enhanced Inulinase production in solid state fermentation by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Journal of Industrial Microbiology & Biotechnology, 2009Co-Authors: Ning Guo, Zhen-ming Chi, Fang Gong, Jun ShengAbstract:In order to isolate Inulinase overproducers of the marine yeast Pichia guilliermondii, strain 1, cells were mutated by using UV light and LiCl2. One mutant (M-30) with enhanced Inulinase production was obtained. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant in solid-state fermentation. The initial moisture, inoculum, the amount ratio of wheat bran to rice bran, temperature, pH for the maximum Inulinase production by the mutant M-30 were found to be 60.5%, 2.5%, 0.42, 30°C and 6.50, respectively. Under the optimized conditions, 455.9 U/grams of dry substrate (gds) of Inulinase activity was reached in the solid state fermentation culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 459.2 U/gds was derived from RSM regression. Under the same conditions, its parent strain only produced 291.0 U/gds of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far.
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Inulinase overproduction by a mutant of the marine yeast pichia guilliermondii using surface response methodology and inulin hydrolysis
Biochemical Engineering Journal, 2009Co-Authors: Ning Guo, Fang Gong, Zhen-ming Chi, Jun Sheng, Zhe ChiAbstract:Abstract In this study, in order to isolate Inulinase overproducers from the marine yeast Pichia guilliermondii, its cells were treated by using UV light and LiCl. The mutant M-30 with enhanced Inulinase production was obtained and was found to be stable after cultivation for 20 generations. Response surface methodology (RSM) was used to optimize the medium compositions and cultivation conditions for Inulinase production by the mutant M-30 in liquid fermentation. Inulin, yeast extract, NaCl, temperature, pH for maximum Inulinase production by the mutant M-30 were found to be 20.0 g/l, 5.0 g/l, 20.0 g/l, 28 °C and 6.5, respectively. Under the optimized conditions, 127.7 U/ml of Inulinase activity was reached in the liquid culture of the mutant M-30 whereas the predicted maximum Inulinase activity of 129.8 U/ml was derived from RSM regression. Under the same conditions, its parent strain only produced 48.1 U/ml of Inulinase activity. This is the highest Inulinase activity produced by the yeast strains reported so far. We also found that inulin could be actively converted into monosaccharides by the crude Inulinase.
