Objective In Corynebacterium crenatum,the adjacent D311 and D312 of N-acetyl-L-glutamate kinase(NAGK),as a key rate-limiting enzyme of L-arginine biosynthesis under substrate regulatory control by arginine,were initia...Objective In Corynebacterium crenatum,the adjacent D311 and D312 of N-acetyl-L-glutamate kinase(NAGK),as a key rate-limiting enzyme of L-arginine biosynthesis under substrate regulatory control by arginine,were initially replaced with two arginine residues to investigate the L-arginine feedback inhibition for NAGK.Methods NAGK enzyme expression was evaluated using a plasmid-based method.Homologous recombination was employed to eliminate the pro B.Results The IC50 and enzyme activity of NAGK M4,in which the D311 R and D312 R amino acid substitutions were combined with the previously reported E19 R and H26 E substitutions,were 3.7-fold and 14.6% higher,respectively,than those of the wild-type NAGK.NAGK M4 was successfully introduced into the C.crenatum MT genome without any genetic markers;the L-arginine yield of C.crenatum MT-M4 was 26.2% higher than that of C.crenatum MT.To further improve upon the L-arginine yield,we constructed the mutant C.crenatum MT-M4 ?pro B.The optimum concentration of L-proline was also investigated in order to determine its contribution to L-arginine yield.After L-proline was added to the medium at 10 mmol/L,the L-arginine yield reached 16.5 g/L after 108 h of shake-flask fermentation,approximately 70.1% higher than the yield attained using C.crenatum MT.Conclusion Feedback inhibition of L-arginine on NAGK in C.crenatum is clearly alleviated by the M4 mutation of NAGK,and deletion of the pro B in C.crenatum from MT to M4 results in a significant increase in arginine production.展开更多
Sulfur emission through fuel combustion is a global problem because it is a major cause of acid rain. Crud oil contains many heterocyclic organic sulfur compounds, among which dibenzothiophene(DBT) and DBTs bearing al...Sulfur emission through fuel combustion is a global problem because it is a major cause of acid rain. Crud oil contains many heterocyclic organic sulfur compounds, among which dibenzothiophene(DBT) and DBTs bearing alkyl substitutions usually are representative compounds. A strain was isolated from refinery sludge and identified as Corynebacterium ZD-1. The behavior of DBT degradation by ZD-1 in aqueous phase was investigated. Corynebacterium ZD-1 could metabolize DBT to 2-hydroxybiphenyl(2-HBP) as the dead-end metabolite through a sulfur-specific pathway. In shake flask culture, ZD-1 had its maximal desulfurization activity in the late exponential growth phase and the specific production rate of 2-HBP was about 0.14(mmol·kg dry cell -1·min -1, mmol·KDC -1·min -1). Active resting cells for desulfurization should be prepared only in this period. 2-HBP inhibited the growth of strain ZD-1, the production of DBT degradation enzymes, and the activity of enzymes. Sulfate inhibited the production of dibenzothiophene(DBT) degradation enzymes but had no effect on the enzymes' activity. The production rates of 2-HBP at lower cell densities were higher and the maximum amount conversion of DBT to 2-HBP(0.067 mmol/L) after 8 h was gained at 9.2 g dry cell/L rather higher cell density. The results indicated that this newly isolated strain could be a promising biocatalyst for DBT desulfurization.展开更多
Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact ar...Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact argR gene from wild-type AS 1.542 was introduced into C. crenatum MT, resulting in C. crenatum MT. sp, and the changes of transcriptional levels of the arginine biosynthetic genes and arginine production were compared between the mutant strain and the recombinant strain. Methods Quantitative real-time polymerase chain reaction was employed to analyze the changes of the related genes at the transcriptional level, electrophoretic mobility shift assays were used to determine ArgR binding with the argCJBDF, argGH, and carAB promoter regions, and arginine production was determined with an automated amino acid analyzer. Results Arginine production assays showed a 69.9% reduction in arginine from 9.01±0.22 mg/mL in C. crenatum MT to 2.71±0.13 mg/mL （P〈0.05） in C. crenatum MT. sp. The argC, argB, argD, argF, argJ, argG, and carA genes were down-regulated significantly in C. crenatum MT. sp compared with those in its parental C. crenatum MT strain. The electrophoretic mobility shift assays showed that the promoter regions were directly bound to the ArgR protein. Conclusion The arginine biosynthetic genes in C crenatum are clearly controlled by the regulator ArgR, and intact ArgR in C. crenatum MT results in a significant descrease in production. negative arginine production.展开更多
The present work aimed to develop a novel strategy to bioremediate the petroleum hydrocarbon contaminants in the environment.Salt tolerant bacterium was isolated from Dagang oilfield,China and identified as Corynebact...The present work aimed to develop a novel strategy to bioremediate the petroleum hydrocarbon contaminants in the environment.Salt tolerant bacterium was isolated from Dagang oilfield,China and identified as Corynebacterium variabile HRJ4 based on 16 S r RNA gene sequence analysis.The bacterium had a high salt tolerant capability and biochar was developed as carrier for the bacterium.The bacteria with biochar were most effective in degradation of n-alkanes（C16,C18,C19,C26,C28） and polycyclic aromatic hydrocarbons（NAP,PYR） mixture.The result demonstrated that immobilization of C.variabile HRJ4 with biochar showed higher degradation of total petroleum hydrocarbons（THPs） up to 78.9%after 7-day of incubation as compared to the free leaving bacteria.The approach of this study will be helpful in clean-up of petroleum-contamination in the environments through bioremediation process using eco-friendly and cost effective materials like biochar.展开更多
The effect of pH of the fermentation medium on cell growth and the production of a novel bioflocculant(named REA-11) by Corynebacterium glutamicum CCTCC M201005 were investigated. The maximum biomass(2.23 g/L) and fl...The effect of pH of the fermentation medium on cell growth and the production of a novel bioflocculant(named REA-11) by Corynebacterium glutamicum CCTCC M201005 were investigated. The maximum biomass(2.23 g/L) and flocculating activity(142.2 U/mL) were simultaneously obtained at the 14th hour when the pH value of the culture medium was maintained at 7.0 during the whole fermentation process. The production of REA-11 kept on a trend of increase till the later phase of fermentation process, which resulted in the ultimate flocculating activity of the culture broth to enhance to nearly 100 U/mL at pH 6.0. A two-stage pH control mode was adopted in REA-11 production in which the pH value of the culture medium was controlled at 7.0 during the first 14 h, then decreased to 6.0 that was maintained until the end of the fermentation process. With the two-stage pH control mode, the maximum flocculating activity reached 178.8 U/mL which was 30% higher than that obtained under the condition of pH 7.0 and the biomass enhanced about 15%. Compared with the fermentation process without pH control, REA-11 production and cell growth via the two-stage pH control mode increased 80% and 25%, respectively.展开更多
Engineered Corynebacterium glutamicum was constructed for L-ornithine production by disrupting genes of argF and proB to prevent the flux away from L-ornithine.Effect of the inactivation of 2-oxoglutarate de-hydrogena...Engineered Corynebacterium glutamicum was constructed for L-ornithine production by disrupting genes of argF and proB to prevent the flux away from L-ornithine.Effect of the inactivation of 2-oxoglutarate de-hydrogenase complex(ODHC) on L-ornithine production was also investigated.It was found that the inactivation of ODHC by knockout of the kgd gene enhanced L-ornithine production.The engineered C.glutamicum ATCC13032(ΔargFΔproBΔkgd) produced L-ornithine up to 4.78 g·L-1 from 0.24 g·L-1 of the wild-type strain.In order to understand the mechanism of L-ornithine production in C.glutamicum ATCC13032(ΔargFΔproBΔkgd) and find out new strategies for further enhancing L-ornithine production,the comparative proteome between the wild-type and the engineered strain was analyzed.L-Ornithine overproduction in the engineered strain was related to the up-regulation of the expression levels of enzymes involved in L-ornithine biosynthesis pathway and down-regulation of the expression levels of proteins involved in pentose phosphate pathway.The overexpression of genes in the upstream pathway of glutamate to increase the availability of endogenous glutamate may further in-crease ornithine production in the engineered C.glutamicum and the ornithine synthesis enzymes(ArgCJBD) may not be the limiting enzymes in the engineered C.glutamicum.展开更多
Fermentation of bioflocculant with Corynebacterium glutamicum was studied by way of kinetic modeling.Lorentzian modified Logistic model, time-corrected Luedeking–Piret and Luedeking–Piret type models were proposed a...Fermentation of bioflocculant with Corynebacterium glutamicum was studied by way of kinetic modeling.Lorentzian modified Logistic model, time-corrected Luedeking–Piret and Luedeking–Piret type models were proposed and applied to describe the cell growth, bioflocculant synthesis and consumption of substrates, with the correlation of initial biomass concentration and initial glucose concentration, respectively. The results showed that these models could well characterize the batch culture process of C. glutamicum at various initial glucose concentrations from 10.0 to 17.5 g·L-1. The initial biomass concentration could shorten the lag time of cell growth,while the maximum biomass concentration was achieved only at the optimal initial glucose concentration of16.22 g·L-1. A novel three-stage fed-batch strategy for bioflocculant production was developed based on the model prediction, in which the lag phase, quick biomass growth and bioflocculant production stages were sequentially proceeded with the adjustment of glucose concentration and dissolved oxygen. Biomass of2.23 g·L-1was obtained and bioflocculant concentration was enhanced to 176.32 mg·L-1, 18.62% and403.63% higher than those in the batch process, respectively, indicating an efficient fed-batch culture strategy for bioflocculant production.展开更多
Objective: Corynebacterium crenatum AS1.542, a Gram-positive bacterium and indigenous nonpatho-genic corynebacteria, is widely exploited for the in-dustrial production of amino acids. The objective of this paper is to...Objective: Corynebacterium crenatum AS1.542, a Gram-positive bacterium and indigenous nonpatho-genic corynebacteria, is widely exploited for the in-dustrial production of amino acids. The objective of this paper is to clarify the genetic information of the arginine biosynthetic pathway, and further more contribute to the improvement of arginine produc-tion. Methods: Polymerase chain reaction (PCR) technology was employed for obtaining the arginine biosynthetic gene sequence, and softwares eg. Laser-gene, BPROM, RNAshapes were used for the analysis of obtained sequences. Results: Arginine biosynethetic gene cluster of C. crenatum, comprising argJ, argB, argD, argF, argR and part of argC, has been ampli-fied and sequenced. The gene order has been estab-lished as argCJBDFR, with a entire length of 6.08kb. Conclusion: An internal promoter was found in the upstream of argB gene, four argBDFR ORFs are lo-cated in a same transcription unit, and the tran-scripiton termination of argC gene is irrelevant with the rho-factor. Comparison with ornithine acetyl-transferase (coded by argJ gene) from C. glutamate, ornithine acetyltransferase from C. crenatum also belongs to the monofunctional enzymes.展开更多
l-Threonine is an important feed additive with the third largest market size among the amino acids produced by microbial fermentation.The GRAS(generally regarded as safe)industrial workhorse Corynebacterium glutamicum...l-Threonine is an important feed additive with the third largest market size among the amino acids produced by microbial fermentation.The GRAS(generally regarded as safe)industrial workhorse Corynebacterium glutamicum is an attractive chassis for l-threonine production.However,the present l-threonine production in C.glutamicum cannot meet the requirement of industrialization due to the relatively low production level of l-threonine and the accumulation of large amounts of by-products(such as l-lysine,l-isoleucine,and glycine).Herein,to enhance the l-threonine biosynthesis in C.glutamicum,releasing the aspartate kinase(LysC)and homoserine dehydrogenase(Hom)from feedback inhibition by l-lysine and l-threonine,respectively,and overexpressing four flux-control genes were performed.Next,to reduce the formation of by-products l-lysine and l-isoleucine without the cause of an auxotrophic phenotype,the feedback regulation of dihydrodipicolinate synthase(DapA)and threonine dehydratase(IlvA)was strengthened by replacing the native enzymes with heterologous analogues with more sensitive feedback inhibition by l-lysine and l-isoleucine,respectively.The resulting strain maintained the capability of synthesizing enough amounts of l-lysine and l-isoleucine for cell biomass formation but exhibited almost no extracellular accumulation of these two amino acids.To further enhance l-threonine production and reduce the by-product glycine,l-threonine exporter and homoserine kinase were overexpressed.Finally,the rationally engineered non-auxotrophic strain ZcglT9 produced 67.63 g/L(17.2%higher)l-threonine with a productivity of 1.20 g/L/h(108.0%higher)in fed-batch fermentation,along with significantly reduced by-product accumulation,representing the record for l-threonine production in C.glutamicum.In this study,we developed a strategy of reconstructing the feedback regulation of amino acid metabolism and successfully applied this strategy to de novo construct a non-auxotrophic l-threonine producing C.glutamicum.The main end by-products including l-lysine,l-isoleucine,and glycine were almost eliminated in fed-batch fermentation of the engineered C.glutamicum strain.This strategy can also be used for engineering producing strains for other amino acids and derivatives.展开更多
Geraniol is a monoterpenoid alcohol with various applications in food,cosmetics,and healthcare.Corynebacterium glutamicum is a potential platform for terpenoids production because it harbors the methylerythritol phosp...Geraniol is a monoterpenoid alcohol with various applications in food,cosmetics,and healthcare.Corynebacterium glutamicum is a potential platform for terpenoids production because it harbors the methylerythritol phosphate pathway.To engineer C.glutamicum to produce geraniol,two different truncated geraniol synthases (GESs) were respectively expressed,and strain LX02 expressing the truncated GESs from Valeriana officinalis (t Vo GES) produced 0.3 mg/L of geraniol.Then,three geranyl diphosphate synthases (GPPSs) were combinatorially co-expressed with t Vo GES to improve geraniol production.The amounts of produced geraniol were all higher than that produced by strain LX02.Strain LX03 co-expressing ERG20 F96W–N127W (ERG20 WW) and t Vo GES produced the highest amount,5.4 mg/L.Subsequently,the co-overexpression of1-deoxy-D-xylulose-5-phosphate synthase (dxs) and isopentenyl diphosphate isomerase (idi) further increased the production to 12.2 mg/L in strain LX03.Lastly,the production of geraniol was increased to 15.2 mg/L via fermentation optimization.To our knowledge,this is the first report on the engineering of C.glutamicum to produce geraniol and thus can serve as a reference for other monoterpenoid production studies.展开更多
[ Objective] This study aimed to investigate the effects of rare earth element lanthanon on the growth and product synthesis of Corynebacterium pekinense. [ Method ] A full factorial experiment was designed to compare...[ Objective] This study aimed to investigate the effects of rare earth element lanthanon on the growth and product synthesis of Corynebacterium pekinense. [ Method ] A full factorial experiment was designed to compare the lysine fermentation of C. pek/nense in medium containing different concentrations of lanthanon. [ Result ] Low doses of lanthanum can promote the growth of C. pekinense and accumulation of metabolite lysine; optimization for distinct components of medium were obtained as 78 mg/L rare earth, 36 g/L glucose, 4.5 g/L beef extract, 20 g/L peptone by Matlab software. [Conclusion] The study initially deter- mined the optimum La＋ additive amount in C. pekinense culture medium and the optimum medium compositions for C. pekinense to produce lysine.展开更多
With the emergence of novel etiologic organisms, panresistance, and invasive medical care infective endocarditis continues to be evasive, requiring newer approaches and modified treatment guidelines. Presented here is...With the emergence of novel etiologic organisms, panresistance, and invasive medical care infective endocarditis continues to be evasive, requiring newer approaches and modified treatment guidelines. Presented here is the case of a 75-year-old male with history of systolic heart failure with an automatic internal cardioverter defibrillator(AICD) implantation and a prosthetic mitral valve who presented with generalized malaise and progressive shortness of breath for 6 d. He was found to have positive blood cultures for gram positive rod shaped bacteria identified as Corynebacterium straitum, but was not considered as the etiological pathogen initially as it a usual skin contaminant. Later this bacterium was found to be the causative agent for the patient's endocarditis. This case highlights the importance of identifying the role of this uncommon commensal in invasive disease. With the use of effective antibiotic regimen and awareness of these new pathogens in invasive disease, mortality and morbidity can be prevented with initiation of early appropriate therapy.展开更多
基金supported by Natural Science Foundation of China,No.31360219 and No.30960012the Open Project Program of Key Laboratory of Functional Small Organic Molecule,Ministry of Education,Jiangxi Normal University(No.KLFS-KF-201414)
文摘Objective In Corynebacterium crenatum,the adjacent D311 and D312 of N-acetyl-L-glutamate kinase(NAGK),as a key rate-limiting enzyme of L-arginine biosynthesis under substrate regulatory control by arginine,were initially replaced with two arginine residues to investigate the L-arginine feedback inhibition for NAGK.Methods NAGK enzyme expression was evaluated using a plasmid-based method.Homologous recombination was employed to eliminate the pro B.Results The IC50 and enzyme activity of NAGK M4,in which the D311 R and D312 R amino acid substitutions were combined with the previously reported E19 R and H26 E substitutions,were 3.7-fold and 14.6% higher,respectively,than those of the wild-type NAGK.NAGK M4 was successfully introduced into the C.crenatum MT genome without any genetic markers;the L-arginine yield of C.crenatum MT-M4 was 26.2% higher than that of C.crenatum MT.To further improve upon the L-arginine yield,we constructed the mutant C.crenatum MT-M4 ?pro B.The optimum concentration of L-proline was also investigated in order to determine its contribution to L-arginine yield.After L-proline was added to the medium at 10 mmol/L,the L-arginine yield reached 16.5 g/L after 108 h of shake-flask fermentation,approximately 70.1% higher than the yield attained using C.crenatum MT.Conclusion Feedback inhibition of L-arginine on NAGK in C.crenatum is clearly alleviated by the M4 mutation of NAGK,and deletion of the pro B in C.crenatum from MT to M4 results in a significant increase in arginine production.
文摘Sulfur emission through fuel combustion is a global problem because it is a major cause of acid rain. Crud oil contains many heterocyclic organic sulfur compounds, among which dibenzothiophene(DBT) and DBTs bearing alkyl substitutions usually are representative compounds. A strain was isolated from refinery sludge and identified as Corynebacterium ZD-1. The behavior of DBT degradation by ZD-1 in aqueous phase was investigated. Corynebacterium ZD-1 could metabolize DBT to 2-hydroxybiphenyl(2-HBP) as the dead-end metabolite through a sulfur-specific pathway. In shake flask culture, ZD-1 had its maximal desulfurization activity in the late exponential growth phase and the specific production rate of 2-HBP was about 0.14(mmol·kg dry cell -1·min -1, mmol·KDC -1·min -1). Active resting cells for desulfurization should be prepared only in this period. 2-HBP inhibited the growth of strain ZD-1, the production of DBT degradation enzymes, and the activity of enzymes. Sulfate inhibited the production of dibenzothiophene(DBT) degradation enzymes but had no effect on the enzymes' activity. The production rates of 2-HBP at lower cell densities were higher and the maximum amount conversion of DBT to 2-HBP(0.067 mmol/L) after 8 h was gained at 9.2 g dry cell/L rather higher cell density. The results indicated that this newly isolated strain could be a promising biocatalyst for DBT desulfurization.
基金supported by Natural Science Foundation of China,No.1360219 and No.30960012
文摘Objective Corynebacterium crenatum MT, a mutant from C. crenatum AS 1.542 with a lethal argR gene, exhibits high arginine production. To confirm the effect of ArgR on arginine biosynthesis in C. crenatum, an intact argR gene from wild-type AS 1.542 was introduced into C. crenatum MT, resulting in C. crenatum MT. sp, and the changes of transcriptional levels of the arginine biosynthetic genes and arginine production were compared between the mutant strain and the recombinant strain. Methods Quantitative real-time polymerase chain reaction was employed to analyze the changes of the related genes at the transcriptional level, electrophoretic mobility shift assays were used to determine ArgR binding with the argCJBDF, argGH, and carAB promoter regions, and arginine production was determined with an automated amino acid analyzer. Results Arginine production assays showed a 69.9% reduction in arginine from 9.01±0.22 mg/mL in C. crenatum MT to 2.71±0.13 mg/mL （P〈0.05） in C. crenatum MT. sp. The argC, argB, argD, argF, argJ, argG, and carA genes were down-regulated significantly in C. crenatum MT. sp compared with those in its parental C. crenatum MT strain. The electrophoretic mobility shift assays showed that the promoter regions were directly bound to the ArgR protein. Conclusion The arginine biosynthetic genes in C crenatum are clearly controlled by the regulator ArgR, and intact ArgR in C. crenatum MT results in a significant descrease in production. negative arginine production.
基金the National Natural Science Foundation of China (No.31270544,41473070)the Hi-Tech Research and Development Program (863) of China (No.2013AA06A205)the 863 achievement transformation program in Tianjin (No.14RCHZSF00144)
文摘The present work aimed to develop a novel strategy to bioremediate the petroleum hydrocarbon contaminants in the environment.Salt tolerant bacterium was isolated from Dagang oilfield,China and identified as Corynebacterium variabile HRJ4 based on 16 S r RNA gene sequence analysis.The bacterium had a high salt tolerant capability and biochar was developed as carrier for the bacterium.The bacteria with biochar were most effective in degradation of n-alkanes（C16,C18,C19,C26,C28） and polycyclic aromatic hydrocarbons（NAP,PYR） mixture.The result demonstrated that immobilization of C.variabile HRJ4 with biochar showed higher degradation of total petroleum hydrocarbons（THPs） up to 78.9%after 7-day of incubation as compared to the free leaving bacteria.The approach of this study will be helpful in clean-up of petroleum-contamination in the environments through bioremediation process using eco-friendly and cost effective materials like biochar.
基金Supported by the Innovative Project for Young Scientific Scholars of Fujian Province(No.2 0 0 2 J0 4 4 )
文摘The effect of pH of the fermentation medium on cell growth and the production of a novel bioflocculant(named REA-11) by Corynebacterium glutamicum CCTCC M201005 were investigated. The maximum biomass(2.23 g/L) and flocculating activity(142.2 U/mL) were simultaneously obtained at the 14th hour when the pH value of the culture medium was maintained at 7.0 during the whole fermentation process. The production of REA-11 kept on a trend of increase till the later phase of fermentation process, which resulted in the ultimate flocculating activity of the culture broth to enhance to nearly 100 U/mL at pH 6.0. A two-stage pH control mode was adopted in REA-11 production in which the pH value of the culture medium was controlled at 7.0 during the first 14 h, then decreased to 6.0 that was maintained until the end of the fermentation process. With the two-stage pH control mode, the maximum flocculating activity reached 178.8 U/mL which was 30% higher than that obtained under the condition of pH 7.0 and the biomass enhanced about 15%. Compared with the fermentation process without pH control, REA-11 production and cell growth via the two-stage pH control mode increased 80% and 25%, respectively.
基金Supported by the National Natural Science Foundation of China (30970089,20876181,20831006)the Natural Science Foundation of Guangdong Province (9351027501000003)
文摘Engineered Corynebacterium glutamicum was constructed for L-ornithine production by disrupting genes of argF and proB to prevent the flux away from L-ornithine.Effect of the inactivation of 2-oxoglutarate de-hydrogenase complex(ODHC) on L-ornithine production was also investigated.It was found that the inactivation of ODHC by knockout of the kgd gene enhanced L-ornithine production.The engineered C.glutamicum ATCC13032(ΔargFΔproBΔkgd) produced L-ornithine up to 4.78 g·L-1 from 0.24 g·L-1 of the wild-type strain.In order to understand the mechanism of L-ornithine production in C.glutamicum ATCC13032(ΔargFΔproBΔkgd) and find out new strategies for further enhancing L-ornithine production,the comparative proteome between the wild-type and the engineered strain was analyzed.L-Ornithine overproduction in the engineered strain was related to the up-regulation of the expression levels of enzymes involved in L-ornithine biosynthesis pathway and down-regulation of the expression levels of proteins involved in pentose phosphate pathway.The overexpression of genes in the upstream pathway of glutamate to increase the availability of endogenous glutamate may further in-crease ornithine production in the engineered C.glutamicum and the ornithine synthesis enzymes(ArgCJBD) may not be the limiting enzymes in the engineered C.glutamicum.
基金Supported by the National Natural Science Foundation of China(21206143,51378444)the program for New Century Excellent Talents of Education Ministry of China(ncet-13-0501)
文摘Fermentation of bioflocculant with Corynebacterium glutamicum was studied by way of kinetic modeling.Lorentzian modified Logistic model, time-corrected Luedeking–Piret and Luedeking–Piret type models were proposed and applied to describe the cell growth, bioflocculant synthesis and consumption of substrates, with the correlation of initial biomass concentration and initial glucose concentration, respectively. The results showed that these models could well characterize the batch culture process of C. glutamicum at various initial glucose concentrations from 10.0 to 17.5 g·L-1. The initial biomass concentration could shorten the lag time of cell growth,while the maximum biomass concentration was achieved only at the optimal initial glucose concentration of16.22 g·L-1. A novel three-stage fed-batch strategy for bioflocculant production was developed based on the model prediction, in which the lag phase, quick biomass growth and bioflocculant production stages were sequentially proceeded with the adjustment of glucose concentration and dissolved oxygen. Biomass of2.23 g·L-1was obtained and bioflocculant concentration was enhanced to 176.32 mg·L-1, 18.62% and403.63% higher than those in the batch process, respectively, indicating an efficient fed-batch culture strategy for bioflocculant production.
文摘Objective: Corynebacterium crenatum AS1.542, a Gram-positive bacterium and indigenous nonpatho-genic corynebacteria, is widely exploited for the in-dustrial production of amino acids. The objective of this paper is to clarify the genetic information of the arginine biosynthetic pathway, and further more contribute to the improvement of arginine produc-tion. Methods: Polymerase chain reaction (PCR) technology was employed for obtaining the arginine biosynthetic gene sequence, and softwares eg. Laser-gene, BPROM, RNAshapes were used for the analysis of obtained sequences. Results: Arginine biosynethetic gene cluster of C. crenatum, comprising argJ, argB, argD, argF, argR and part of argC, has been ampli-fied and sequenced. The gene order has been estab-lished as argCJBDFR, with a entire length of 6.08kb. Conclusion: An internal promoter was found in the upstream of argB gene, four argBDFR ORFs are lo-cated in a same transcription unit, and the tran-scripiton termination of argC gene is irrelevant with the rho-factor. Comparison with ornithine acetyl-transferase (coded by argJ gene) from C. glutamate, ornithine acetyltransferase from C. crenatum also belongs to the monofunctional enzymes.
基金funded by the National Key Research and Development Program of China(2021YFC2100900)the Tianjin Synthetic Biotechnology Innovation Capacity Improvement Project(TSBICIP-CXRC-058)+2 种基金the National Natural Science Foundation of China(32270101)the Key R&D Program of Shandong Province(2021CXGC010602)the Youth Innovation Promotion Association of Chinese Academy of Sciences(2021177)。
文摘l-Threonine is an important feed additive with the third largest market size among the amino acids produced by microbial fermentation.The GRAS(generally regarded as safe)industrial workhorse Corynebacterium glutamicum is an attractive chassis for l-threonine production.However,the present l-threonine production in C.glutamicum cannot meet the requirement of industrialization due to the relatively low production level of l-threonine and the accumulation of large amounts of by-products(such as l-lysine,l-isoleucine,and glycine).Herein,to enhance the l-threonine biosynthesis in C.glutamicum,releasing the aspartate kinase(LysC)and homoserine dehydrogenase(Hom)from feedback inhibition by l-lysine and l-threonine,respectively,and overexpressing four flux-control genes were performed.Next,to reduce the formation of by-products l-lysine and l-isoleucine without the cause of an auxotrophic phenotype,the feedback regulation of dihydrodipicolinate synthase(DapA)and threonine dehydratase(IlvA)was strengthened by replacing the native enzymes with heterologous analogues with more sensitive feedback inhibition by l-lysine and l-isoleucine,respectively.The resulting strain maintained the capability of synthesizing enough amounts of l-lysine and l-isoleucine for cell biomass formation but exhibited almost no extracellular accumulation of these two amino acids.To further enhance l-threonine production and reduce the by-product glycine,l-threonine exporter and homoserine kinase were overexpressed.Finally,the rationally engineered non-auxotrophic strain ZcglT9 produced 67.63 g/L(17.2%higher)l-threonine with a productivity of 1.20 g/L/h(108.0%higher)in fed-batch fermentation,along with significantly reduced by-product accumulation,representing the record for l-threonine production in C.glutamicum.In this study,we developed a strategy of reconstructing the feedback regulation of amino acid metabolism and successfully applied this strategy to de novo construct a non-auxotrophic l-threonine producing C.glutamicum.The main end by-products including l-lysine,l-isoleucine,and glycine were almost eliminated in fed-batch fermentation of the engineered C.glutamicum strain.This strategy can also be used for engineering producing strains for other amino acids and derivatives.
基金supported by the National Natural Science Foundation of China (No. 21878220)。
文摘Geraniol is a monoterpenoid alcohol with various applications in food,cosmetics,and healthcare.Corynebacterium glutamicum is a potential platform for terpenoids production because it harbors the methylerythritol phosphate pathway.To engineer C.glutamicum to produce geraniol,two different truncated geraniol synthases (GESs) were respectively expressed,and strain LX02 expressing the truncated GESs from Valeriana officinalis (t Vo GES) produced 0.3 mg/L of geraniol.Then,three geranyl diphosphate synthases (GPPSs) were combinatorially co-expressed with t Vo GES to improve geraniol production.The amounts of produced geraniol were all higher than that produced by strain LX02.Strain LX03 co-expressing ERG20 F96W–N127W (ERG20 WW) and t Vo GES produced the highest amount,5.4 mg/L.Subsequently,the co-overexpression of1-deoxy-D-xylulose-5-phosphate synthase (dxs) and isopentenyl diphosphate isomerase (idi) further increased the production to 12.2 mg/L in strain LX03.Lastly,the production of geraniol was increased to 15.2 mg/L via fermentation optimization.To our knowledge,this is the first report on the engineering of C.glutamicum to produce geraniol and thus can serve as a reference for other monoterpenoid production studies.
基金Supported by Tianjin Vocational Institute Education Program ( 20061004)
文摘[ Objective] This study aimed to investigate the effects of rare earth element lanthanon on the growth and product synthesis of Corynebacterium pekinense. [ Method ] A full factorial experiment was designed to compare the lysine fermentation of C. pek/nense in medium containing different concentrations of lanthanon. [ Result ] Low doses of lanthanum can promote the growth of C. pekinense and accumulation of metabolite lysine; optimization for distinct components of medium were obtained as 78 mg/L rare earth, 36 g/L glucose, 4.5 g/L beef extract, 20 g/L peptone by Matlab software. [Conclusion] The study initially deter- mined the optimum La＋ additive amount in C. pekinense culture medium and the optimum medium compositions for C. pekinense to produce lysine.
文摘With the emergence of novel etiologic organisms, panresistance, and invasive medical care infective endocarditis continues to be evasive, requiring newer approaches and modified treatment guidelines. Presented here is the case of a 75-year-old male with history of systolic heart failure with an automatic internal cardioverter defibrillator(AICD) implantation and a prosthetic mitral valve who presented with generalized malaise and progressive shortness of breath for 6 d. He was found to have positive blood cultures for gram positive rod shaped bacteria identified as Corynebacterium straitum, but was not considered as the etiological pathogen initially as it a usual skin contaminant. Later this bacterium was found to be the causative agent for the patient's endocarditis. This case highlights the importance of identifying the role of this uncommon commensal in invasive disease. With the use of effective antibiotic regimen and awareness of these new pathogens in invasive disease, mortality and morbidity can be prevented with initiation of early appropriate therapy.
