Use of slow glucose feeding as supporting carbon source in lactose autoinduction medium improves the robustness of protein expression at different aeration conditions

Abstract This study aimed to evaluate the growth, respiratory activity, and biodegradation of chlorpyrifos in cultures of Azotobacter vinelandii ATCC 12837. A strategy based on the modification of culture media and aeration conditions was carried out to increase the cell concentration of A. vinelandii , in order to favor and determine its tolerance to chlorpyrifos and its degradation ability. The culture in shaken flasks, using sucrose as a carbon source, significantly improved the growth compared to media with mannitol. When the strain was cultivated under oxygen-limited (5.5, 11.25 mmol L- 1 h- 1 ) and no-oxygen-limited conditions (22 mmol L -1 h -1 ), the growth parameters were not affected. In cultures in a liquid medium with chlorpyrifos, the bacteria tolerated a high pesticide concentration (500 ppm) and the growth parameters were improved even under conditions with a reduced carbon source (sucrose 2 g L -1 ). The strain degraded 99.6 % of chlorpyrifos at 60 h of cultivation, in co-metabolism with sucrose; notably, A. vinelandii ATCC 12837 reduced by 50% the initial pesticide concentration in only 6 h (DT 50 ).

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Optimization of carbon source and glucose feeding strategy for improvement of L-isoleucine production byEscherichia coli

Biotechnology & Biotechnological Equipment ◽

10.1080/13102818.2015.1006899 ◽

2015 ◽

Vol 29 (2) ◽

pp. 374-380 ◽

Cited By ~ 11

Author(s):

Jian Wang ◽

Bing Wen ◽

Qingyang Xu ◽

Xixian Xie ◽

Ning Chen

Keyword(s):

Carbon Source ◽

Feeding Strategy ◽

Glucose Feeding ◽

Byescherichia Coli

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Growth, respiratory activity and chlorpyrifos biodegradation in cultures of Azotobacter vinelandii ATCC 12837

AMB Express ◽

10.1186/s13568-021-01339-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Victoria Conde-Avila ◽

Carlos Peña ◽

Beatriz Pérez-Armendáriz ◽

Octavio Loera ◽

Carmen Martínez Valenzuela ◽

...

Keyword(s):

Carbon Source ◽

Liquid Medium ◽

Respiratory Activity ◽

Azotobacter Vinelandii ◽

Cell Concentration ◽

Culture Media ◽

Growth Parameters ◽

Pesticide Concentration ◽

Aeration Conditions ◽

Degradation Ability

AbstractThis study aimed to evaluate the growth, respiratory activity, and biodegradation of chlorpyrifos in cultures of Azotobacter vinelandii ATCC 12837. A strategy based on the modification of culture media and aeration conditions was carried out to increase the cell concentration of A. vinelandii, in order to favor and determine its tolerance to chlorpyrifos and its degradation ability. The culture in shaken flasks, using sucrose as a carbon source, significantly improved the growth compared to media with mannitol. When the strain was cultivated under oxygen-limited (5.5, 11.25 mmol L−1 h−1) and no-oxygen-limited conditions (22 mmol L−1 h−1), the growth parameters were not affected. In cultures in a liquid medium with chlorpyrifos, the bacteria tolerated a high pesticide concentration (500 ppm) and the growth parameters were improved even under conditions with a reduced carbon source (sucrose 2 g L−1). The strain degraded 99.6% of chlorpyrifos at 60 h of cultivation, in co-metabolism with sucrose; notably, A. vinelandii ATCC 12837 reduced by 50% the initial pesticide concentration in only 6 h (DT50). Graphical Abstract

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A Novel Regulated Hybrid Promoter That Permits Autoinduction of Heterologous Protein Expression inKluyveromyces lactis

Applied and Environmental Microbiology ◽

10.1128/aem.00542-19 ◽

2019 ◽

Vol 85 (14) ◽

Cited By ~ 2

Author(s):

Hassan Sakhtah ◽

Juliane Behler ◽

Alana Ali-Reynolds ◽

Thomas B. Causey ◽

Saulius Vainauskas ◽

...

Keyword(s):

Gene Expression ◽

Carbon Source ◽

Protein Expression ◽

Recombinant Proteins ◽

Heterologous Protein ◽

Kluyveromyces Lactis ◽

Heterologous Protein Expression ◽

Heterologous Proteins ◽

Content Type ◽

Hybrid Promoter

ABSTRACTThe yeastKluyveromyces lactishas been a successful host for the production of heterologous proteins for over 30 years. Currently, the galactose-/lactose-inducible and glucose-repressibleLAC4promoter (PLAC4) is the most widely used promoter to drive recombinant protein expression inK. lactis. However, PLAC4is not fully repressed in the presence of glucose and significant protein expression still occurs. Thus, PLAC4is not suitable in processes where tight regulation of heterologous gene expression is required. In this study, we devised a novelK. lactispromoter system that is both strong and tightly controllable. We first tested several different endogenousK. lactispromoters for their ability to express recombinant proteins. A novel hybrid promoter (termed P350) was created by combining segments of twoK. lactispromoters, namely, the strong constitutive PGAP1promoter and the carbon source-sensitive PICL1promoter. We demonstrate that P350is tightly repressed in the presence of glucose or glycerol and becomes derepressed upon depletion of these compounds by the growing cells. We further illustrate the utility of P350-controlled protein expression in shake flask and high-cell-density bioreactor cultivation strategies. The P350hybrid promoter is a strong derepressible promoter for use in autoinduction of one-step fermentation processes for the production of heterologous proteins inK. lactis.IMPORTANCEThe yeastKluyveromyces lactisis an important host for the expression of recombinant proteins at both laboratory and industrial scales. However, the system lacks a tightly regulated promoter that permits controlled expression of heterologous proteins. In this study, we report the engineering of a highly regulated strong hybrid promoter (termed P350) for use inK. lactis. P350is tightly repressed by glucose or glycerol in the medium but strongly promotes gene expression once the carbon source has been consumed by the cells. This feature permits heterologous protein expression to be “autoinduced” at any scale without the addition of a gratuitous inducer molecule or changing feed solutions.

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Glutamate dehydrogenase regulation in callus cultures of Nicotiana plumbaginifolia: effect of glucose feeding and carbon source starvation on the isoenzymatic pattern

Plant Cell & Environment ◽

10.1111/j.1365-3040.1991.tb01533.x ◽

1991 ◽

Vol 14 (6) ◽

pp. 613-618 ◽

Cited By ~ 15

Author(s):

E. MAESTRI ◽

F. M. RESTIVO ◽

M. GULLI ◽

F. TASSI

Keyword(s):

Carbon Source ◽

Glutamate Dehydrogenase ◽

Nicotiana Plumbaginifolia ◽

Callus Cultures ◽

Glucose Feeding ◽

Effect Of Glucose ◽

Isoenzymatic Pattern

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Morphological Characterization of Mycobacteriophage R1

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100051281 ◽

1974 ◽

Vol 32 ◽

pp. 254-255

Author(s):

B. L. Soloff ◽

T. A. Rado

Keyword(s):

Carbon Source ◽

Stationary Phase ◽

Growth Phase ◽

Minimal Medium ◽

Morphological Characterization ◽

Logarithmic Growth Phase ◽

Complete Lysis ◽

Lysogenic Culture ◽

Logarithmic Growth

Mycobacteriophage R1 was originally isolated from a lysogenic culture of M. butyricum. The virus was propagated on a leucine-requiring derivative of M. smegmatis, 607 leu−, isolated by nitrosoguanidine mutagenesis of typestrain ATCC 607. Growth was accomplished in a minimal medium containing glycerol and glucose as carbon source and enriched by the addition of 80 μg/ ml L-leucine. Bacteria in early logarithmic growth phase were infected with virus at a multiplicity of 5, and incubated with aeration for 8 hours. The partially lysed suspension was diluted 1:10 in growth medium and incubated for a further 8 hours. This permitted stationary phase cells to re-enter logarithmic growth and resulted in complete lysis of the culture.

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