scholarly journals PII Signal Transduction Protein GlnK Alleviates Feedback Inhibition of N-Acetyl-L-Glutamate Kinase by L-Arginine in Corynebacterium glutamicum

2020 ◽  
Vol 86 (8) ◽  
Author(s):  
Meijuan Xu ◽  
Mi Tang ◽  
Jiamin Chen ◽  
Taowei Yang ◽  
Xian Zhang ◽  
...  
Keyword(s):  
Signal Transduction ◽  
Amino Acid ◽  
Glutamic Acid ◽  
Corynebacterium Glutamicum ◽  
Feedback Inhibition ◽  
Amino Acid Biosynthesis ◽  
Arginine Biosynthesis ◽  
Acid Biosynthesis ◽  
Content Type ◽  
Pii Protein

ABSTRACT PII signal transduction proteins are ubiquitous and highly conserved in bacteria, archaea, and plants and play key roles in controlling nitrogen metabolism. However, research on biological functions and regulatory targets of PII proteins remains limited. Here, we illustrated experimentally that the PII protein Corynebacterium glutamicum GlnK (CgGlnK) increased l-arginine yield when glnK was overexpressed in Corynebacterium glutamicum. Data showed that CgGlnK regulated l-arginine biosynthesis by upregulating the expression of genes of the l-arginine metabolic pathway and interacting with N-acetyl-l-glutamate kinase (CgNAGK), the rate-limiting enzyme in l-arginine biosynthesis. Further assays indicated that CgGlnK contributed to alleviation of the feedback inhibition of CgNAGK caused by l-arginine. In silico analysis of the binding interface of CgGlnK-CgNAGK suggested that the B and T loops of CgGlnK mainly interacted with C and N domains of CgNAGK. Moreover, F11, R47, and K85 of CgGlnK were identified as crucial binding sites that interact with CgNAGK via hydrophobic interaction and H bonds, and these interactions probably had a positive effect on maintaining the stability of the complex. Collectively, this study reveals PII-NAGK interaction in nonphotosynthetic microorganisms and further provides insights into the regulatory mechanism of PII on amino acid biosynthesis in corynebacteria. IMPORTANCE Corynebacteria are safe industrial producers of diverse amino acids, including l-glutamic acid and l-arginine. In this study, we showed that PII protein GlnK played an important role in l-glutamic acid and l-arginine biosynthesis in C. glutamicum. Through clarifying the molecular mechanism of CgGlnK in l-arginine biosynthesis, the novel interaction between CgGlnK and CgNAGK was revealed. The alleviation of l-arginine inhibition of CgNAGK reached approximately 48.21% by CgGlnK addition, and the semi-inhibition constant of CgNAGK increased 1.4-fold. Furthermore, overexpression of glnK in a high-yield l-arginine-producing strain and fermentation of the recombinant strain in a 5-liter bioreactor led to a remarkably increased production of l-arginine, 49.978 g/liter, which was about 22.61% higher than that of the initial strain. In conclusion, this study provides a new strategy for modifying amino acid biosynthesis in C. glutamicum.

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 Microbial Origin of Plant-Type 2-Keto-3-Deoxy-d-arabino-Heptulosonate 7-Phosphate Synthases, Exemplified by the Chorismate- and Tryptophan-Regulated Enzyme from Xanthomonas campestris

2001 ◽  
Vol 183 (13) ◽  
pp. 4061-4070 ◽  
Author(s):  
Guillermo Gosset ◽  
Carol A. Bonner ◽  
Roy A. Jensen
Keyword(s):  
Amino Acid ◽  
Aromatic Amino Acid ◽  
Xanthomonas Campestris ◽  
Higher Plants ◽  
Feedback Inhibition ◽  
Amino Acid Biosynthesis ◽  
Partial Purification ◽  
Initial Reaction ◽  
Acid Biosynthesis ◽  
Aromatic Amino Acid Biosynthesis

ABSTRACT Enzymes performing the initial reaction of aromatic amino acid biosynthesis, 2-keto-3-deoxy-d-arabino-heptulosonate 7-phosphate (DAHP) synthases, exist as two distinct homology classes. The three classic Escherichia coli paralogs are AroAI proteins, but many members of theBacteria possess the AroAII class of enzyme, sometimes in combination with AroAI proteins. AroAII DAHP synthases until now have been shown to be specifically dedicated to secondary metabolism (e.g., formation of ansamycin antibiotics or phenazine pigment). In contrast, here we show that the Xanthomonas campestris AroAII protein functions as the sole DAHP synthase supporting aromatic amino acid biosynthesis. X. campestris AroAII was cloned in E. coli by functional complementation, and genes corresponding to two possible translation starts were expressed. We developed a 1-day partial purification method (>99%) for the unstable protein. The recombinant AroAII protein was found to be subject to an allosteric pattern of sequential feedback inhibition in which chorismate is the prime allosteric effector.l-Tryptophan was found to be a minor feedback inhibitor. An N-terminal region of 111 amino acids may be located in the periplasm since a probable inner membrane-spanning region is predicted. Unlike chloroplast-localized AroAII of higher plants, X. campestris AroAII was not hysteretically activated by dithiols. Compared to plant AroAII proteins, differences in divalent metal activation were also observed. Phylogenetic tree analysis shows that AroAII originated within theBacteria domain, and it seems probable that higher-plant plastids acquired AroAII from a gram-negative bacterium via endosymbiosis. The X. campestris AroAII protein is suggested to exemplify a case of analog displacement whereby an ancestral aroA I species was discarded, with thearoA II replacement providing an alternative pattern of allosteric control. Three subgroups of AroAIIproteins can be recognized: a large, central group containing the plant enzymes and that from X. campestris, one defined by a three-residue deletion near the conserved KPRS motif, and one possessing a larger deletion further downstream.

scholarly journals Identification of Genes That Confer Sediment Fitness to Desulfovibrio desulfuricans G20

2007 ◽  
Vol 73 (19) ◽  
pp. 6305-6312 ◽  
Author(s):  
Qingwei Luo ◽  
Jennifer L. Groh ◽  
Jimmy D. Ballard ◽  
Lee R. Krumholz
Keyword(s):  
Signal Transduction ◽  
Dna Repair ◽  
Amino Acid ◽  
Desulfovibrio Desulfuricans ◽  
Amino Acid Biosynthesis ◽  
Hydrogenase Activity ◽  
Functional Categories ◽  
Acid Biosynthesis ◽  
Genes Encoding ◽  
Insertion Elements

ABSTRACT Signature-tagged mutants of Desulfovibrio desulfuricans G20 were screened, and 97 genes crucial for sediment fitness were identified. These genes belong to functional categories including signal transduction, binding and transport, insertion elements, and others. Mutants with mutations in genes encoding proteins involved in amino acid biosynthesis, hydrogenase activity, and DNA repair were further characterized.

scholarly journals GapMind: Automated Annotation of Amino Acid Biosynthesis

mSystems ◽  
2020 ◽  
Vol 5 (3) ◽  
Author(s):  
Morgan N. Price ◽  
Adam M. Deutschbauer ◽  
Adam P. Arkin
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Building Blocks ◽  
Amino Acid Biosynthesis ◽  
Acid Biosynthesis ◽  
Content Type ◽  
Alternative Pathways ◽  
Automated Annotation ◽  
A Genome ◽  
Minimal Media

ABSTRACT GapMind is a Web-based tool for annotating amino acid biosynthesis in bacteria and archaea (http://papers.genomics.lbl.gov/gaps). GapMind incorporates many variant pathways and 130 different reactions, and it analyzes a genome in just 15 s. To avoid error-prone transitive annotations, GapMind relies primarily on a database of experimentally characterized proteins. GapMind correctly handles fusion proteins and split proteins, which often cause errors for best-hit approaches. To improve GapMind’s coverage, we examined genetic data from 35 bacteria that grow in defined media without amino acids, and we filled many gaps in amino acid biosynthesis pathways. For example, we identified additional genes for arginine synthesis with succinylated intermediates in Bacteroides thetaiotaomicron, and we propose that Dyella japonica synthesizes tyrosine from phenylalanine. Nevertheless, for many bacteria and archaea that grow in minimal media, genes for some steps still cannot be identified. To help interpret potential gaps, GapMind checks if they match known gaps in related microbes that can grow in minimal media. GapMind should aid the identification of microbial growth requirements. IMPORTANCE Many microbes can make all of the amino acids (the building blocks of proteins). In principle, we should be able to predict which amino acids a microbe can make, and which it requires as nutrients, by checking its genome sequence for all of the necessary genes. However, in practice, it is difficult to check for all of the alternative pathways. Furthermore, new pathways and enzymes are still being discovered. We built an automated tool, GapMind, to annotate amino acid biosynthesis in bacterial and archaeal genomes. We used GapMind to list gaps: cases where a microbe makes an amino acid but a complete pathway cannot be identified in its genome. We used these gaps, together with data from mutants, to identify new pathways and enzymes. However, for most bacteria and archaea, we still do not know how they can make all of the amino acids.

scholarly journals Allosteric Feedback Inhibition Enables Robust Amino Acid Biosynthesis in E. coli by Enforcing Enzyme Overabundance

Cell Systems ◽  
2019 ◽  
Vol 8 (1) ◽  
pp. 66-75.e8 ◽  
Author(s):  
Timur Sander ◽  
Niklas Farke ◽  
Christoph Diehl ◽  
Michelle Kuntz ◽  
Timo Glatter ◽  
...  
Keyword(s):  
Amino Acid ◽  
Feedback Inhibition ◽  
Amino Acid Biosynthesis ◽  
Acid Biosynthesis ◽  
E Coli
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