High-level expression of the methanol-inducible β-galactosidase gene by perfusion culture of recombinant Pichia pastoris using a shaken ceramic membrane flask

Ohashi, Ryo; Mochizuki, Eiko; Kamoshita, Yuya; Suzuki, Takahiro

doi:10.1016/s0922-338x(98)80032-9

Cited by 10 publications

(5 citation statements)

References 7 publications

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“…With the addition of 10% peptone or 0.2 mol/l of arginine to the culture medium, the fibrinolytic activity of prourokinaseannnexin V reached 7.8 kU/ml at 168 h of culture, which was more than 10-fold that obtained without the supplement in P. pastoris [18]. In addition, Polypepton was used as the feed medium for effective b-galactosidase production in a perfusion culture from P. pastoris using a shaker ceramic membrane flask and a two-stage fed-batch culture [19]. Ammonia sulfate was also used for preventing the sereprotease degradation of recombinant human serum albumin secreted by P. pastoris during culture [20].…”

Section: Discussionmentioning

confidence: 95%

Enhanced production of mouse α-amylase by feeding combined nitrogen and carbon sources in fed-batch culture of recombinant Pichia pastoris

Choi¹,

Park

2006

Process Biochemistry

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The optimal culture conditions for attaining high levels of expression of mouse a-amylase in the methylotrophic yeast Pichia pastoris were investigated. Cell growth and enzyme production were found to be optimal at a temperature of 28 8C and a pH of 6.0. Yeast extract and peptone were the most appropriate nitrogen sources for the expression of a-amylase. When a mixture of 20 g/l peptone and 20 g/l yeast extract was used as the nitrogen source, a-amylase activity reached 300 U/ml. The expression of a-amylase in a fed-batch culture using methanol as the sole carbon source reached 720 U/ml at 3 days of culture. However, when a methanol-to-glycerol mixture was used at ratio of 1:0.5 at a feed rate of 3 g/h, the maximum a-amylase activity was 1.14 kU/ml. Under the optimal culture conditions, when 40 g/l of a mixture of yeast extract and peptone (1:1) was added to the culture at 3 and 5 days the final a-amylase activity increased to 2.4 kU/ml, which was 5.8-and 2.1-fold higher than the activity in the batch culture and in the fed-batch culture with feeding a mixture of methanol and glycerol at a ratio of 1:0.5, respectively. This study demonstrates that an addition of the appropriate amount of an organic nitrogen source to the fed-batch culture is effective to improve high level of expression of mouse a-amylase in the methylotrophic yeast Pichia pastoris. #

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Section: Discussionmentioning

confidence: 95%

Enhanced production of mouse α-amylase by feeding combined nitrogen and carbon sources in fed-batch culture of recombinant Pichia pastoris

Choi¹,

Park

2006

Process Biochemistry

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“…Therefore, the mutant may have some potential applications in food and pharmaceutical industries. However, the recombinant β-galactosidase activity produced by Pichia pastoris GS115 carrying the β-galactosidase gene by the feeding of methanol reached 152 U/ml at 168 h [18].…”

Section: Discussionmentioning

confidence: 99%

Enhanced β-Galactosidase Production from Whey Powder by a Mutant of the Psychrotolerant Yeast Guehomyces pullulans 17–1 for Hydrolysis of Lactose

Zhao²,

Wang

et al. 2011

Appl Biochem Biotechnol

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In order to isolate β-galactosidase overproducers of the psychrotolerant yeast Guehomyces pullulans 17-1, its cells were mutated by using nitrosoguanidine (NTG). One mutant (NTG-133) with enhanced β-galactosidase production was obtained. The mutant grown in the production medium with 30.0 g/l lactose and 2.0 g/l glucose could produce more β-galactosidase than the same mutant grown in the production medium with only 30.0 g/l lactose while β-galactosidase production by its wild type was sensitive to the presence of glucose in the medium. It was found that 40.0 g/l of the whey powder was the most suitable for β-galactosidase production by the mutant. After optimization of the medium and cultivation conditions, the mutant could produce 29.2 U/ml of total β-galactosidase activity within 132 h at the flask level while the mutant could produce 48.1 U/ml of total β-galactosidase activity within 144 h in 2-l fermentor. Over 77.1% of lactose in the whey powder (5.0% w/v) was hydrolyzed in the presence of the β-galactosidase activity of 280 U/g of lactose within 9 h while over 77.0% of lactose in the whey was hydrolyzed in the presence of β-galactosidase activity of 280 U/g of lactose within 6 h. This was the first time to show that the β-galactosidase produced by the psychrotolerant yeast could be used for hydrolysis of lactose in the whey powder and whey.

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“…Other more complex continuous schemes than chemostats and nutriostats have also been reported. Perfusion culture technique using a shaken ceramic membrane flask (SCM flask) has been applied to the recombinant production of β-galactosidase [ 88 , 89 ]. A rotary membrane separation system was used in a continuous fermentation with cell recycling to obtain high cell concentration and high thrombomodulin production [ 49 ].…”

Section: Reviewmentioning

confidence: 99%

Operational strategies, monitoring and control of heterologous protein production in the methylotrophic yeast Pichia pastoris under different promoters: A review

et al. 2006

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The methylotrophic yeast Pichia pastoris has been widely reported as a suitable expression system for heterologous protein production. The use of different phenotypes under PAOX promoter, other alternative promoters, culture medium, and operational strategies with the objective to maximize either yield or productivity of the heterologous protein, but also to obtain a repetitive product batch to batch to get a robust process for the final industrial application have been reported. Medium composition, kinetics growth, fermentation operational strategies from fedbatch to continuous cultures using different phenotypes with the most common PAOX promoter and other novel promoters (GAP, FLD, ICL), the use of mixed substrates, on-line monitoring of the key fermentation parameters (methanol) and control algorithms applied to the bioprocess are reviewed and discussed in detail.

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High-level expression of the methanol-inducible β-galactosidase gene by perfusion culture of recombinant Pichia pastoris using a shaken ceramic membrane flask

Cited by 10 publications

References 7 publications

Enhanced production of mouse α-amylase by feeding combined nitrogen and carbon sources in fed-batch culture of recombinant Pichia pastoris

Enhanced production of mouse α-amylase by feeding combined nitrogen and carbon sources in fed-batch culture of recombinant Pichia pastoris

Enhanced β-Galactosidase Production from Whey Powder by a Mutant of the Psychrotolerant Yeast Guehomyces pullulans 17–1 for Hydrolysis of Lactose

Operational strategies, monitoring and control of heterologous protein production in the methylotrophic yeast Pichia pastoris under different promoters: A review

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