scholarly journals Impact of Classical Strain Improvement of Penicillium rubens on Amino Acid Metabolism during β-Lactam Production

2019 ◽  
Vol 86 (3) ◽  
Author(s):  
Min Wu ◽  
Ciprian G. Crismaru ◽  
Oleksandr Salo ◽  
Roel A. L. Bovenberg ◽  
Arnold J. M. Driessen
Keyword(s):  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Strain Improvement ◽  
Tryptophan Synthase ◽  
Cysteine Biosynthesis ◽  
Penicillin Production ◽  
Content Type ◽  
Classical Strain ◽  
The Impact

ABSTRACT To produce high levels of β-lactams, the filamentous fungus Penicillium rubens (previously named Penicillium chrysogenum) has been subjected to an extensive classical strain improvement (CSI) program during the last few decades. This has led to the accumulation of many mutations that were spread over the genome. Detailed analysis reveals that several mutations targeted genes that encode enzymes involved in amino acid metabolism, in particular biosynthesis of l-cysteine, one of the amino acids used for β-lactam production. To examine the impact of the mutations on enzyme function, the respective genes with and without the mutations were cloned and expressed in Escherichia coli, purified, and enzymatically analyzed. Mutations severely impaired the activities of a threonine and serine deaminase, and this inactivates metabolic pathways that compete for l-cysteine biosynthesis. Tryptophan synthase, which converts l-serine into l-tryptophan, was inactivated by a mutation, whereas a mutation in 5-aminolevulinate synthase, which utilizes glycine, was without an effect. Importantly, CSI caused increased expression levels of a set of genes directly involved in cysteine biosynthesis. These results suggest that CSI has resulted in improved cysteine biosynthesis by the inactivation of the enzymatic conversions that directly compete for resources with the cysteine biosynthetic pathway, consistent with the notion that cysteine is a key component during penicillin production. IMPORTANCE Penicillium rubens is an important industrial producer of β-lactam antibiotics. High levels of penicillin production were enforced through extensive mutagenesis during a classical strain improvement (CSI) program over 70 years. Several mutations targeted amino acid metabolism and resulted in enhanced l-cysteine biosynthesis. This work provides a molecular explanation for the interrelation between secondary metabolite production and amino acid metabolism and how classical strain improvement has resulted in improved production strains.

scholarly journals Conserved Pyridoxal 5'-Phosphate-Binding Protein YggS Impacts Amino Acid Metabolism through Pyridoxine 5'-Phosphate inEscherichia coli

2019 ◽  
Vol 85 (11) ◽  
Author(s):  
Tomokazu Ito ◽  
Kana Yamamoto ◽  
Ran Hori ◽  
Ayako Yamauchi ◽  
Diana M. Downs ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Biosynthetic Pathway ◽  
Acid Metabolism ◽  
Binding Protein ◽  
Protein Family ◽  
Pleiotropic Effects ◽  
Vitamin B ◽  
Content Type ◽  
E Coli

ABSTRACTEscherichia coliYggS (COG0325) is a member of the highly conserved pyridoxal 5′-phosphate (PLP)-binding protein (PLPBP) family. Recent studies suggested a role for this protein family in the homeostasis of vitamin B6and amino acids. The deletion or mutation of a member of this protein family causes pleiotropic effects in many organisms and is causative of vitamin B6-dependent epilepsy in humans. To date, little has been known about the mechanism by which lack of YggS results in these diverse phenotypes. In this study, we determined that the pyridoxine (PN) sensitivity observed inyggS-deficientE. coliwas caused by the pyridoxine 5′-phosphate (PNP)-dependent overproduction of Val, which is toxic toE. coli. The data suggest that theyggSmutation impacts Val accumulation by perturbing the biosynthetic of Thr from homoserine (Hse). Exogenous Hse inhibited the growth of theyggSmutant, caused further accumulation of PNP, and increased the levels of some intermediates in the Thr-Ile-Val metabolic pathways. Blocking the Thr biosynthetic pathway or decreasing the intracellular PNP levels abolished the perturbations of amino acid metabolism caused by the exogenous PN and Hse. Our data showed that a high concentration of intracellular PNP is the root cause of at least some of the pleiotropic phenotypes described for ayggSmutant ofE. coli.IMPORTANCERecent studies showed that deletion or mutation of members of the YggS protein family causes pleiotropic effects in many organisms. Little is known about the causes, mechanisms, and consequences of these diverse phenotypes. It was previously shown thatyggSmutations inE. coliresult in the accumulation of PNP and some metabolites in the Ile/Val biosynthetic pathway. This work revealed that some exogenous stresses increase the aberrant accumulation of PNP in theyggSmutant. In addition, the current report provides evidence indicating that some, but not all, of the phenotypes of theyggSmutant inE. coliare due to the elevated PNP level. These results will contribute to continuing efforts to determine the molecular functions of the members of the YggS protein family.

scholarly journals Trophic cooperation promotes bacterial survival of Staphylococcus aureus and Pseudomonas aeruginosa

2020 ◽  
Author(s):  
Laura Camus ◽  
Paul Briaud ◽  
Sylvère Bastien ◽  
Sylvie Elsen ◽  
Anne Doléans-Jordheim ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Metabolic Process ◽  
Bacterial Survival ◽  
Beneficial Effects ◽  
The Impact

AbstractIn the context of infection, Pseudomonas aeruginosa and Staphylococcus aureus are frequently co-isolated, particularly in cystic fibrosis (CF) patients. Within lungs, the two pathogens exhibit a range of competitive and coexisting interactions. In the present study, we explored the impact of S. aureus on the physiology of P. aeruginosa in the context of coexistence. Transcriptomic analyses showed that S. aureus significantly and specifically affects the expression of numerous genes involved in P. aeruginosa carbon and amino acid metabolism. In particular, 65% of the strains presented considerable overexpression of the genes involved in the acetoin catabolic (aco) pathway. We demonstrated that acetoin is (i) produced by clinical S. aureus strains, (ii) detected in sputa from CF patients, and (iii) involved in P. aeruginosa’s aco system induction. Furthermore, acetoin is catabolized by P. aeruginosa, a metabolic process that improves the survival of both pathogens by providing a new carbon source for P. aeruginosa and avoiding the toxic accumulation of acetoin on S. aureus. Due to its beneficial effects on both bacteria, acetoin catabolism could testify to the establishment of trophic cooperation between S. aureus and P. aeruginosa in the CF lung environment, thus promoting their persistence.

scholarly journals Suppression of a Thermosensitive zipA Cell Division Mutant by Altering Amino Acid Metabolism

2017 ◽  
Vol 200 (2) ◽  
Author(s):  
Daniel E. Vega ◽  
William Margolin
Keyword(s):  
Escherichia Coli ◽  
Cell Division ◽  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Cytoplasmic Membrane ◽  
Amino Acid Biosynthesis ◽  
Acid Biosynthesis ◽  
Content Type ◽  
Cell Division Protein

ABSTRACTZipA is essential for cell division inEscherichia coli, acting early in the process to anchor polymers of FtsZ to the cytoplasmic membrane. Along with FtsA, FtsZ and ZipA form a proto-ring at midcell that recruits additional proteins to eventually build the division septum. Cells carrying the thermosensitivezipA1allele divide fairly normally at 30°C in rich medium but cease dividing at temperatures above 34°C, forming long filaments. In a search for suppressors of thezipA1allele, we found that deletions of specific genes involved in amino acid biosynthesis could partially rescue cell growth and division at 34°C or 37°C but not at 42°C. Notably, although a diverse group of amino acid biosynthesis gene deletions could partially rescue the growth ofzipA1cells at 34°C, only deletions of genes related to the biosynthesis of threonine, glycine, serine, and methionine could rescue growth at 37°C. Adding exogenous pyridoxal 5-phosphate (PLP), a cofactor for many of the enzymes affected by this study, partially suppressedzipA1mutant thermosensitivity. For many of the deletions, PLP had an additive rescuing effect on thezipA1mutant. Moreover, added PLP partially suppressed the thermosensitivity offtsQandftsKmutants and weakly suppressed anftsImutant, but it failed to suppressftsAorftsZthermosensitive mutants. Along with the ability of a deletion ofmetCto partially suppress theftsKmutant, our results suggest that perturbations of amino acid metabolic pathways, particularly those that redirect the flow of carbon away from the synthesis of threonine, glycine, or methionine, are able to partially rescue some cell division defects.IMPORTANCECell division of bacteria, such asEscherichia coli, is essential for their successful colonization. It is becoming increasingly clear that nutritional status and central metabolism can affect bacterial size and shape; for example, a metabolic enzyme (OpgH) can moonlight as a regulator of FtsZ, an essential cell division protein. Here, we demonstrate a link between amino acid metabolism and ZipA, another essential cell division protein that binds directly to FtsZ and tethers it to the cytoplasmic membrane. Our evidence suggests that altering flux through the methionine-threonine-glycine-serine pathways and supplementing with the enzyme cofactor pyridoxal-5-phosphate can partially compensate for an otherwise lethal defect in ZipA, as well as several other cell division proteins.

scholarly journals Sulfur Amino Acid Metabolism in Pregnancy: The Impact of Methionine in the Maternal Diet

2006 ◽  
Vol 136 (6) ◽  
pp. 1701S-1705S ◽  
Author(s):  
William D. Rees ◽  
Fiona A. Wilson ◽  
Christopher A. Maloney
Keyword(s):  
Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Maternal Diet ◽  
Sulfur Amino Acid ◽  
The Impact ◽  
In Pregnancy

scholarly journals Putative Horizontally Acquired Genes, Highly Transcribed during Yersinia pestis Flea Infection, Are Induced by Hyperosmotic Stress and Function in Aromatic Amino Acid Metabolism

2020 ◽  
Vol 202 (11) ◽  
Author(s):  
Luary C. Martínez-Chavarría ◽  
Janelle Sagawa ◽  
Jessica Irons ◽  
Angela K. Hinz ◽  
Athena Lemon ◽  
...  
Keyword(s):  
Gene Expression ◽  
Amino Acid ◽  
Yersinia Pestis ◽  
Salinity Stress ◽  
Aromatic Amino Acid ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Gc Content ◽  
Content Type ◽  
Aromatic Amino Acid Metabolism

ABSTRACT While alternating between insects and mammals during its life cycle, Yersinia pestis, the flea-transmitted bacterium that causes plague, regulates its gene expression appropriately to adapt to these two physiologically disparate host environments. In fleas competent to transmit Y. pestis, low-GC-content genes y3555, y3551, and y3550 are highly transcribed, suggesting that these genes have a highly prioritized role in flea infection. Here, we demonstrate that y3555, y3551, and y3550 are transcribed as part of a single polycistronic mRNA comprising the y3555, y3554, y3553, y355x, y3551, and y3550 genes. Additionally, y355x-y3551-y3550 compose another operon, while y3550 can be also transcribed as a monocistronic mRNA. The expression of these genes is induced by hyperosmotic salinity stress, which serves as an explicit environmental stimulus that initiates transcriptional activity from the predicted y3550 promoter. Y3555 has homology to pyridoxal 5′-phosphate (PLP)-dependent aromatic aminotransferases, while Y3550 and Y3551 are homologous to the Rid protein superfamily (YjgF/YER057c/UK114) members that forestall damage caused by reactive intermediates formed during PLP-dependent enzymatic activity. We demonstrate that y3551 specifically encodes an archetypal RidA protein with 2-aminoacrylate deaminase activity but Y3550 lacks Rid deaminase function. Heterologous expression of y3555 generates a critical aspartate requirement in a Salmonella enterica aspC mutant, while its in vitro expression, and specifically its heterologous coexpression with y3550, enhances the growth rate of an Escherichia coli ΔaspC ΔtyrB mutant in a defined minimal amino acid-supplemented medium. Our data suggest that the y3555, y3551, and y3550 genes operate cooperatively to optimize aromatic amino acid metabolism and are induced under conditions of hyperosmotic salinity stress. IMPORTANCE Distinct gene repertoires are expressed during Y. pestis infection of its flea and mammalian hosts. The functions of many of these genes remain predicted or unknown, necessitating their characterization, as this may provide a better understanding of Y. pestis specialized biological adaptations to the discrete environments of its two hosts. This study provides functional context to adjacently clustered horizontally acquired genes predominantly expressed in the flea host by deciphering their fundamental processes with regard to (i) transcriptional organization, (ii) transcription activation signals, and (iii) biochemical function. Our data support a role for these genes in osmoadaptation and aromatic amino acid metabolism, highlighting these as preferential processes by which Y. pestis gene expression is modulated during flea infection.

scholarly journals Sink/Source Balance of Leaves Influences Amino Acid Pools and Their Associated Metabolic Fluxes in Winter Oilseed Rape (Brassica napus L.)

Metabolites ◽  
2020 ◽  
Vol 10 (4) ◽  
pp. 150 ◽  
Author(s):  
Younès Dellero ◽  
Maud Heuillet ◽  
Nathalie Marnet ◽  
Floriant Bellvert ◽  
Pierre Millard ◽  
...  
Keyword(s):  
Amino Acid ◽  
Oilseed Rape ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
De Novo ◽  
Vegetative Stage ◽  
Primary Metabolism ◽  
Winter Oilseed Rape ◽  
Brassica Napus L ◽  
The Impact

Nitrogen remobilization processes from source to sink tissues in plants are determinant for seed yield and their implementation results in a complete reorganization of the primary metabolism during sink/source transition. Here, we decided to characterize the impact of the sink/source balance on amino acid metabolism in the leaves of winter oilseed rape grown at the vegetative stage. We combined a quantitative metabolomics approach with an instationary 15N-labeling experiment by using [15N]L-glycine as a metabolic probe on leaf ranks with a gradual increase in their source status. We showed that the acquisition of the source status by leaves was specifically accompanied by a decrease in asparagine, glutamine, proline and S-methyl-l-cysteine sulphoxide contents and an increase in valine and threonine contents. Dynamic analysis of 15N enrichment and concentration of amino acids revealed gradual changes in the dynamics of amino acid metabolism with respect to the sink/source status of leaf ranks. Notably, nitrogen assimilation into valine, threonine and proline were all decreased in source leaves compared to sink leaves. Overall, our results suggested a reduction in de novo amino acid biosynthesis during sink/source transition at the vegetative stage.

scholarly journals Global effects of homocysteine on transcription in Escherichia coli: induction of the gene for the major cold-shock protein, CspA

Microbiology ◽  
2006 ◽  
Vol 152 (8) ◽  
pp. 2221-2231 ◽  
Author(s):  
Katy R. Fraser ◽  
Nina L. Tuite ◽  
Arvind Bhagwat ◽  
Conor P. O'Byrne
Keyword(s):  
Escherichia Coli ◽  
Amino Acid ◽  
Growth Medium ◽  
Amino Acid Metabolism ◽  
Cold Shock ◽  
Acid Metabolism ◽  
Cold Shock Protein ◽  
Rt Pcr ◽  
Life Threatening ◽  
The Impact

Homocysteine (Hcy) is a thiol-containing amino acid that is considered to be medically important because it is linked to the development of several life-threatening diseases in humans, including cardiovascular disease and stroke. It inhibits the growth of Escherichia coli when supplied in the growth medium. Growth inhibition is believed to arise as a result of partial starvation for isoleucine, which occurs because Hcy perturbs the biosynthesis of this amino acid. This study attempted to further elucidate the inhibitory mode of action of Hcy by examining the impact of exogenously supplied Hcy on the transcriptome. Using gene macroarrays the transcript levels corresponding to 68 genes were found to be reproducibly altered in the presence of 0.5 mM Hcy. Of these genes, the biggest functional groups affected were those involved in translation (25 genes) and in amino acid metabolism (19 genes). Genes involved in protection against oxidative stress were repressed in Hcy-treated cells and this correlated with a decrease in catalase activity. The gene showing the strongest induction by Hcy was cspA, which encodes the major cold-shock protein CspA. RT-PCR and reporter fusion experiments confirmed that cspA was induced by Hcy. Induction of cspA by Hcy was not caused by nutritional upshift, a stimulus known to induce CspA expression, nor was it dependent on the presence of a functional CspA protein. The induction of cspA by Hcy was suppressed when isoleucine was included in the growth medium. These data suggest that the induction of CspA expression in the presence of Hcy occurs because of a limitation for isoleucine. The possibility that Hcy-induced cspA expression is triggered by translational stalling that occurs when the cells are limited for isoleucine is discussed.

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