scholarly journals Genes of theN-Methylglutamate Pathway Are Essential for Growth of Methylobacterium extorquens DM4 with Monomethylamine

2014 ◽  
Vol 80 (11) ◽  
pp. 3541-3550 ◽  
Author(s):  
Christelle Gruffaz ◽  
Emilie E. L. Muller ◽  
Yousra Louhichi-Jelail ◽  
Yella R. Nelli ◽  
Gilles Guichard ◽  
...  
Keyword(s):  
Carbon Source ◽  
Nitrogen Source ◽  
Gene Clusters ◽  
Sole Source ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Methylotrophic Bacteria ◽  
Methylobacterium Extorquens ◽  
Content Type ◽  
Carbon And Nitrogen ◽  
In The Wild

ABSTRACTMonomethylamine (MMA, CH3NH2) can be used as a carbon and nitrogen source by many methylotrophic bacteria.Methylobacterium extorquensDM4 lacks the MMA dehydrogenase encoded bymaugenes, which inM. extorquensAM1 is essential for growth on MMA. Identification and characterization of minitransposon mutants with an MMA-dependent phenotype showed that strain DM4 grows with MMA as the sole source of carbon, energy, and nitrogen by theN-methylglutamate (NMG) pathway. Independent mutations were found in a chromosomal region containing the genesgmaS,mgsABC, andmgdABCDfor the three enzymes of the pathway, γ-glutamylmethylamide (GMA) synthetase, NMG synthase, and NMG dehydrogenase, respectively. Reverse transcription-PCR confirmed the operonic structure of the two divergent gene clustersmgsABC-gmaSandmgdABCDand their induction during growth with MMA. The genesmgdABCDandmgsABCwere found to be essential for utilization of MMA as a carbon and nitrogen source. The genegmaSwas essential for MMA utilization as a carbon source, but residual growth of mutant DM4gmaSgrowing with succinate and MMA as a nitrogen source was observed. Plasmid copies ofgmaSand thegmaShomolog METDI4690, which encodes a protein 39% identical to GMA synthetase, fully restored the ability of mutants DM4gmaSand DM4gmaSΔmetdi4690 to use MMA as a carbon and nitrogen source. Similarly, chemically synthesized GMA, the product of GMA synthetase, could be used as a nitrogen source for growth in the wild-type strain, as well as in DM4gmaSand DM4gmaSΔmetdi4690 mutants. The NADH:ubiquinone oxidoreductase respiratory complex component NuoG was also found to be essential for growth with MMA as a carbon source.

scholarly journals Key Enzymes Enabling the Growth of Arthrobacter sp. Strain JBH1 with Nitroglycerin as the Sole Source of Carbon and Nitrogen

2012 ◽  
Vol 78 (10) ◽  
pp. 3649-3655 ◽  
Author(s):  
Johana Husserl ◽  
Joseph B. Hughes ◽  
Jim C. Spain
Keyword(s):  
Heterologous Expression ◽  
Nitrogen Source ◽  
Sole Source ◽  
Glycerol Kinase ◽  
Enzyme Assays ◽  
Content Type ◽  
Key Enzymes ◽  
Carbon And Nitrogen ◽  
Subsequent Transformation

ABSTRACTFlavoprotein reductases that catalyze the transformation of nitroglycerin (NG) to dinitro- or mononitroglycerols enable bacteria containing such enzymes to use NG as the nitrogen source. The inability to use the resulting mononitroglycerols limits most strains to incomplete denitration of NG. Recently,Arthrobacterstrain JBH1 was isolated for the ability to grow on NG as the sole source of carbon and nitrogen, but the enzymes and mechanisms involved were not established. Here, the enzymes that enable theArthrobacterstrain to incorporate NG into a productive pathway were identified. Enzyme assays indicated that the transformation of nitroglycerin to mononitroglycerol is NADPH dependent and that the subsequent transformation of mononitroglycerol is ATP dependent. Cloning and heterologous expression revealed that a flavoprotein catalyzes selective denitration of NG to 1-mononitroglycerol (1-MNG) and that 1-MNG is transformed to 1-nitro-3-phosphoglycerol by a glycerol kinase homolog. Phosphorylation of the nitroester intermediate enables the subsequent denitration of 1-MNG in a productive pathway that supports the growth of the isolate and mineralization of NG.

scholarly journals Carbaryl as a Carbon and Nitrogen Source: an Inducible Methylamine Metabolic Pathway at the Biochemical and Molecular Levels inPseudomonassp. Strain C5pp

2018 ◽  
Vol 84 (24) ◽  
Author(s):  
Kamini ◽  
Rakesh Sharma ◽  
Narayan S. Punekar ◽  
Prashant S. Phale
Keyword(s):  
Carbon Source ◽  
Nitrogen Source ◽  
Sole Source ◽  
Biogeochemical Cycles ◽  
Ecological Systems ◽  
Suitable Candidate ◽  
Carbon And Nitrogen ◽  
Metabolic Versatility ◽  
Soil Isolate ◽  
Water Ecosystems

ABSTRACTCarbaryl is the most widely used carbamate family pesticide, and its persistent nature causes it to pollute both soil and water ecosystems. Microbes maintain the Earth’s biogeochemical cycles by metabolizing various compounds present in the matter, including xenobiotics, as a sole source of carbon, nitrogen, and energy. Soil isolatePseudomonassp. strain C5pp metabolizes carbaryl efficiently as the carbon source. Periplasmic carbaryl hydrolase catalyzes the conversion of carbaryl to 1-naphthol and methylamine. 1-Naphthol was further used as a carbon source via gentisate, whereas the metabolic fate of methylamine is not known. Here, we demonstrate that strain C5pp showed efficient growth on carbaryl when supplied as a carbon and nitrogen source, suggesting that the methylamine generated was used as the nitrogen source. Genes involved in the methylamine metabolism were annotated and characterized at the biochemical and molecular level. Transcriptional and enzyme activity studies corroborate that the γ-glutamylmethylamide/N-methylglutamate (GMA/NMG) pathway is involved in the metabolism of carbaryl and methylamine as a nitrogen source. Compared to carbaryl, methylamine was found to be an effective inducer for the metabolic and transporter genes. Strain C5pp also harbored genes involved in sarcosine metabolism that were cotranscribed and induced by sarcosine. The presence of inducible pathways for metabolism of carbaryl as a nitrogen and carbon source helps in complete and efficient mineralization of carbaryl by strain C5pp, thereby maintaining the biogeochemical cycles.IMPORTANCEThe degradation of xenobiotics plays a significant role in the environment to maintain ecological systems as well as to prevent the imbalance of biogeochemical cycles via carbon-nitrogen cycling. Carbaryl is the most widely used pesticide from the carbamate family.Pseudomonassp. strain C5pp, capable of utilizing carbaryl as a carbon and nitrogen source for its growth, subsequently helps in complete remediation of carbaryl. Thus, it maintains the ecosystem by balancing the biogeochemical cycles. The metabolic versatility and genetic diversity of strain C5pp for the transformation of contaminants like carbaryl and 1-naphthol into less harmful products make it a suitable candidate from the perspective of bioremediation.

scholarly journals Production of Feruloyl Esterase from Aspergillus niger by Solid-State Fermentation on Different Carbon Sources

2011 ◽  
Vol 2011 ◽  
pp. 1-4 ◽  
Author(s):  
Shiyi Ou ◽  
Jing Zhang ◽  
Yong Wang ◽  
Ning Zhang
Keyword(s):  
Carbon Source ◽  
Nitrogen Source ◽  
Wheat Bran ◽  
Cell Walls ◽  
Carbon Sources ◽  
Feruloyl Esterase ◽  
Plant Cell Walls ◽  
Optimal Conditions ◽  
Carbon And Nitrogen ◽  
Maize Bran

A mixture of wheat bran with maize bran as a carbon source and addition of (NH4)SO4 as nitrogen source was found to significantly increase production of feruloyl esterase (FAE) enzyme compared with wheat bran as a sole carbon and nitrogen source. The optimal conditions in conical flasks were carbon source (30 g) to water 1 : 1, maize bran to wheat bran 1 : 2, (NH4)SO4 1.2 g and MgSO4 70 mg. Under these conditions, FAE activity was 7.68 mU/g. The FAE activity on the mixed carbon sources showed, high activity against the plant cell walls contained in the cultures.

scholarly journals A Mannose Family Phosphotransferase System Permease and Associated Enzymes Are Required for Utilization of Fructoselysine and Glucoselysine in Salmonella enterica Serovar Typhimurium

2015 ◽  
Vol 197 (17) ◽  
pp. 2831-2839 ◽  
Author(s):  
Katherine A. Miller ◽  
Robert S. Phillips ◽  
Paul B. Kilgore ◽  
Grady L. Smith ◽  
Timothy R. Hoover
Keyword(s):  
Nitrogen Source ◽  
Nitrogen Sources ◽  
Maillard Reaction Products ◽  
Reaction Products ◽  
Phosphotransferase System ◽  
Carbon And Nitrogen Sources ◽  
Content Type ◽  
Carbon And Nitrogen ◽  
Serovar Typhimurium ◽  
Food Borne Illness

ABSTRACTSalmonella entericserovar Typhimurium, a major cause of food-borne illness, is capable of using a variety of carbon and nitrogen sources. Fructoselysine and glucoselysine are Maillard reaction products formed by the reaction of glucose or fructose, respectively, with the ε-amine group of lysine. We report here thatS. Typhimurium utilizes fructoselysine and glucoselysine as carbon and nitrogen sources via a mannose family phosphotransferase (PTS) encoded bygfrABCD(glucoselysine/fructoselysine PTS components EIIA, EIIB, EIIC, and EIID; locus numbers STM14_5449 to STM14_5454 inS. Typhimurium 14028s). Genes coding for two predicted deglycases within thegfroperon,gfrEandgfrF, were required for growth with glucoselysine and fructoselysine, respectively. GfrF demonstrated fructoselysine-6-phosphate deglycase activity in a coupled enzyme assay. The biochemical and genetic analyses were consistent with a pathway in which fructoselysine and glucoselysine are phosphorylated at the C-6 position of the sugar by the GfrABCD PTS as they are transported across the membrane. The resulting fructoselysine-6-phosphate and glucoselysine-6-phosphate subsequently are cleaved by GfrF and GfrE to form lysine and glucose-6-phosphate or fructose-6-phosphate. Interestingly, althoughS. Typhimurium can use lysine derived from fructoselysine or glucoselysine as a sole nitrogen source, it cannot use exogenous lysine as a nitrogen source to support growth. Expression ofgfrABCDEFwas dependent on the alternative sigma factor RpoN (σ54) and an RpoN-dependent LevR-like activator, which we designated GfrR.IMPORTANCESalmonellaphysiology has been studied intensively, but there is much we do not know regarding the repertoire of nutrients these bacteria are able to use for growth. This study shows that a previously uncharacterized PTS and associated enzymes function together to transport and catabolize fructoselysine and glucoselysine. Knowledge of the range of nutrients thatSalmonellautilizes is important, as it could lead to the development of new strategies for reducing the load ofSalmonellain food animals, thereby mitigating its entry into the human food supply.

scholarly journals Molecular Mechanism ofN,N-Dimethylformamide Degradation inMethylobacteriumsp. Strain DM1

2019 ◽  
Vol 85 (12) ◽  
Author(s):  
Xinyu Lu ◽  
Weiwei Wang ◽  
Lige Zhang ◽  
Haiyang Hu ◽  
Ping Xu ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Gene Clusters ◽  
Sole Source ◽  
Human Beings ◽  
Sequencing Data ◽  
Carbon And Nitrogen ◽  
Redundant Genes ◽  
Evolutionary Advantage ◽  
Redundant Gene ◽  
Phenotype Identification

ABSTRACTN,N-Dimethylformamide (DMF) is one of the most common xenobiotic chemicals, and it can be easily emitted into the environment, where it causes harm to human beings. Herein, an efficient DMF-degrading strain, DM1, was isolated and identified asMethylobacteriumsp. This strain can use DMF as the sole source of carbon and nitrogen. Whole-genome sequencing of strain DM1 revealed that it has a 5.66-Mbp chromosome and a 200-kbp megaplasmid. The plasmid pLVM1 specifically harbors the genes essential for the initial steps of DMF degradation, and the chromosome carries the genes facilitating subsequent methylotrophic metabolism. Through analysis of the transcriptome sequencing data, the complete mineralization pathway and redundant gene clusters of DMF degradation were elucidated. The dimethylformamidase (DMFase) gene was heterologously expressed, and DMFase was purified and characterized. Plasmid pLVM1 is catabolically crucial for DMF utilization, as evidenced by the phenotype identification of the plasmid-free strain. This study systematically elucidates the molecular mechanisms of DMF degradation byMethylobacterium.IMPORTANCEDMF is a hazardous pollutant that has been used in the chemical industry, pharmaceutical manufacturing, and agriculture. Biodegradation as a method for removing DMF has received increasing attention. Here, we identified an efficient DMF degrader,Methylobacteriumsp. strain DM1, and characterized the complete DMF mineralization pathway and enzymatic properties of DMFase in this strain. This study provides insights into the molecular mechanisms and evolutionary advantage of DMF degradation facilitated by plasmid pLVM1 and redundant genes in strain DM1, suggesting the emergence of new ecotypes ofMethylobacterium.

scholarly journals The Gene Cluster forpara-Nitrophenol Catabolism Is Responsible for 2-Chloro-4-Nitrophenol Degradation in Burkholderia sp. Strain SJ98

2014 ◽  
Vol 80 (19) ◽  
pp. 6212-6222 ◽  
Author(s):  
Jun Min ◽  
Jun-Jie Zhang ◽  
Ning-Yi Zhou
Keyword(s):  
Gene Cluster ◽  
Gene Knockout ◽  
Gene Clusters ◽  
Sole Source ◽  
Pcr Analysis ◽  
Content Type ◽  
Reverse Transcription Pcr ◽  
Biochemical Analyses ◽  
Nitrophenol Degradation

ABSTRACTBurkholderiasp. strain SJ98 (DSM 23195) utilizes 2-chloro-4-nitrophenol (2C4NP) orpara-nitrophenol (PNP) as a sole source of carbon and energy. Here, by genetic and biochemical analyses, a 2C4NP catabolic pathway different from those of all other 2C4NP utilizers was identified with chloro-1,4-benzoquinone (CBQ) as an intermediate. Reverse transcription-PCR analysis showed that all of thepnpgenes in thepnpABA1CDEFcluster were located in a single operon, which is significantly different from the genetic organization of all other previously reported PNP degradation gene clusters, in which the structural genes were located in three different operons. All of the Pnp proteins were purified to homogeneity as His-tagged proteins. PnpA, a PNP 4-monooxygenase, was found to be able to catalyze the monooxygenation of 2C4NP to CBQ. PnpB, a 1,4-benzoquinone reductase, has the ability to catalyze the reduction of CBQ to chlorohydroquinone. Moreover, PnpB is also able to enhance PnpA activityin vitroin the conversion of 2C4NP to CBQ. Genetic analyses indicated thatpnpAplays an essential role in the degradation of both 2C4NP and PNP by gene knockout and complementation. In addition to being responsible for the lower pathway of PNP catabolism, PnpCD, PnpE, and PnpF were also found to be likely involved in that of 2C4NP catabolism. These results indicated that the catabolism of 2C4NP and that of PNP share the same gene cluster in strain SJ98. These findings fill a gap in our understanding of the microbial degradation of 2C4NP at the molecular and biochemical levels.

scholarly journals Multiple and Interconnected Pathways for l-Lysine Catabolism in Pseudomonas putida KT2440

2005 ◽  
Vol 187 (21) ◽  
pp. 7500-7510 ◽  
Author(s):  
Olga Revelles ◽  
Manuel Espinosa-Urgel ◽  
Tobias Fuhrer ◽  
Uwe Sauer ◽  
Juan L. Ramos
Keyword(s):  
Pseudomonas Putida ◽  
Nitrogen Source ◽  
Pseudomonas Putida Kt2440 ◽  
Carbon And Nitrogen ◽  
New Genes ◽  
In The Wild ◽  
Isogenic Mutants ◽  
Hydrolysis Of ◽  
Successful Colonization ◽  
Lysine Catabolism

ABSTRACT l-Lysine catabolism in Pseudomonas putida KT2440 was generally thought to occur via the aminovalerate pathway. In this study we demonstrate the operation of the alternative aminoadipate pathway with the intermediates d-lysine, l-pipecolate, and aminoadipate. The simultaneous operation of both pathways for the use of l-lysine as the sole carbon and nitrogen source was confirmed genetically. Mutants with mutations in either pathway failed to use l-lysine as the sole carbon and nitrogen source, although they still used l-lysine as the nitrogen source, albeit at reduced growth rates. New genes were identified in both pathways, including the davB and davA genes that encode the enzymes involved in the oxidation of l-lysine to δ-aminovaleramide and the hydrolysis of the latter to δ-aminovalerate, respectively. The amaA, dkpA, and amaB genes, in contrast, encode proteins involved in the transformation of Δ1-piperidine-2-carboxylate into aminoadipate. Based on l-[U-13C, U-15N]lysine experiments, we quantified the relative use of pathways in the wild type and its isogenic mutants. The fate of 13C label of l-lysine indicates that in addition to the existing connection between the d- and l-lysine pathways at the early steps of the catabolism of l-lysine mediated by a lysine racemase, there is yet another interconnection at the lower end of the pathways in which aminoadipate is channeled to yield glutarate. This study establishes an unequivocal relationship between gene and pathway enzymes in the metabolism of l-lysine, which is of crucial importance for the successful colonization of the rhizosphere of plants by this microorganism.

Octopine and octopinic acid utilization in a nonfluorescent Pseudomonas sp.: enhancement by spontaneous mutation and lack of effect from curing of a plasmid

1987 ◽  
Vol 33 (6) ◽  
pp. 534-540 ◽  
Author(s):  
Rodolphe Boivin ◽  
Hélène Lebeuf ◽  
Patrice Dion
Keyword(s):  
Carbon Source ◽  
Nitrogen Source ◽  
Spontaneous Mutation ◽  
Pseudomonas Sp ◽  
Wild Type ◽  
Carbon And Nitrogen ◽  
Mutant Derivative ◽  
Alternative Carbon Source ◽  
Derivatives Of ◽  
Derivative Strain

An octopine-utilizing bacterium isolated from soil, named strain 92, was identified to the genus level as a nonfluorescent Pseudomonas. Utilization of octopine by strain 92 depended upon the presence of an alternative carbon source, while octopinic acid was not utilized. A mutant derivative, strain RB100, acquired simultaneously the capacity to utilize octopine as the sole carbon and nitrogen source and octopinic acid as the nitrogen source. The mutation rate was estimated to be about 10−8 per cell per generation. Both the wild type and mutant carried the 68.5-kilobase cryptic plasmid pDLB 1. When the plasmid present in strain RB100 was labelled with Tn5 and transferred to strain 92, the phenotype of the recipient was not modified with respect to octopine utilization, suggesting that this phenotype was not determined by pDLB1. This suggestion was confirmed by using unstable derivatives of pDLB1 to cure strain RB100 and showing that the cured strain was not affected in its capacity to utilize opines.

scholarly journals Discovery of 16-Demethylrifamycins by Removing the Predominant Polyketide Biosynthesis Pathway in Micromonospora sp. Strain TP-A0468

2018 ◽  
Vol 85 (4) ◽  
Author(s):  
Qiang Zhou ◽  
Guang-Cai Luo ◽  
Huizhan Zhang ◽  
Gong-Li Tang
Keyword(s):  
Natural Products ◽  
Transcriptional Activation ◽  
Metabolic Flux ◽  
Genome Mining ◽  
Gene Clusters ◽  
Bioinformatic Analysis ◽  
Transcriptional Analysis ◽  
Content Type ◽  
In The Wild ◽  
New Compounds

ABSTRACT A number of strategies have been developed to mine novel natural products based on biosynthetic gene clusters and there have been dozens of successful cases facilitated by the development of genomic sequencing. During our study on biosynthesis of the antitumor polyketide kosinostatin (KST), we found that the genome of Micromonospora sp. strain TP-A0468, the producer of KST, contains other potential polyketide gene clusters, with no encoded products detected. Deletion of kst cluster led to abolishment of KST and the enrichment of several new compounds, which were isolated and characterized as 16-demethylrifamycins (referred to here as compounds 3 to 6). Transcriptional analysis demonstrated that the expression of the essential genes related to the biosynthesis of compounds 3 to 6 was comparable to the level in the wild-type and in the kst cluster deletion strain. This indicates that the accumulation of these compounds was due to the redirection of metabolic flux rather than transcriptional activation. Genetic disruption, chemical complementation, and bioinformatic analysis revealed that the production of compounds 3 to 6 was accomplished by cross talk between the two distantly placed polyketide gene clusters pks3 and M-rif. This finding not only enriches the analogue pool and the biosynthetic diversity of rifamycins but also provides an auxiliary strategy for natural product discovery through genome mining in polyketide-producing microorganisms. IMPORTANCE Natural products are essential in the development of novel clinically used drugs. Discovering new natural products and modifying known compounds are still the two main ways to generate new candidates. Here, we have discovered several rifamycins with varied skeleton structures by redirecting the metabolic flux from the predominant polyketide biosynthetic pathway to the rifamycin pathway in the marine actinomycetes species Micromonospora sp. strain TP-A0468. Rifamycins are indispensable chemotherapeutics in the treatment of various diseases such as tuberculosis, leprosy, and AIDS-related mycobacterial infections. This study exemplifies a useful method for the discovery of cryptic natural products in genome-sequenced microbes. Moreover, the 16-demethylrifamycins and their genetically manipulable producer provide a new opportunity in the construction of novel rifamycin derivates to aid in the defense against the ever-growing drug resistance of Mycobacterium tuberculosis.

scholarly journals Lactobacillus casei Ferments theN-Acetylglucosamine Moiety of Fucosyl-α-1,3-N-Acetylglucosamine and Excretes l-Fucose

2012 ◽  
Vol 78 (13) ◽  
pp. 4613-4619 ◽  
Author(s):  
Jesús Rodríguez-Díaz ◽  
Antonio Rubio-del-Campo ◽  
María J. Yebra
Keyword(s):  
Carbon Source ◽  
Lactobacillus Casei ◽  
Genomic Sequence ◽  
Protein A ◽  
Physiological Role ◽  
Gene Clusters ◽  
Transcriptional Analysis ◽  
Phosphotransferase System ◽  
Content Type

ABSTRACTWe have previously characterized fromLactobacillus caseiBL23 three α-l-fucosidases, AlfA, AlfB, and AlfC, which hydrolyzein vitronatural fucosyl-oligosaccharides. In this work, we have shown thatL. caseiis able to grow in the presence of fucosyl-α-1,3-N-acetylglucosamine (Fuc-α-1,3-GlcNAc) as a carbon source. Interestingly,L. caseiexcretes thel-fucose moiety during growth on Fuc-α-1,3-GlcNAc, indicating that only theN-acetylglucosamine moiety is being metabolized. Analysis of the genomic sequence ofL. caseiBL23 shows that downstream fromalfB, which encodes the α-l-fucosidase AlfB, a gene,alfR, that encodes a transcriptional regulator is present. Divergently fromalfB, three genes,alfEFG, that encode proteins with homology to the enzyme IIAB (EIIAB), EIIC, and EIID components of a mannose-class phosphoenolpyruvate:sugar phosphotransferase system (PTS) are present. Inactivation of eitheralfBoralfFabolishes the growth ofL. caseion Fuc-α-1,3-GlcNAc. This proves that AlfB is involved in Fuc-α-1,3-GlcNAc metabolism and that the transporter encoded byalfEFGparticipates in the uptake of this disaccharide. A mutation in the PTS general component enzyme I does not eliminate the utilization of Fuc-α-1,3-GlcNAc, suggesting that the transport via the PTS encoded byalfEFGis not coupled to phosphorylation of the disaccharide. Transcriptional analysis withalfRandccpAmutants shows that the two gene clustersalfBRandalfEFGare regulated by substrate-specific induction mediated by the inactivation of the transcriptional repressor AlfR and by carbon catabolite repression mediated by the catabolite control protein A (CcpA). This work reports for the first time the characterization of the physiological role of an α-l-fucosidase in lactic acid bacteria and the utilization of Fuc-α-1,3-GlcNAc as a carbon source for bacteria.

Sign in / Sign up

Export Citation Format

Share Document