scholarly journals YjeH Is a Novel Exporter of l-Methionine and Branched-Chain Amino Acids in Escherichia coli

2015 ◽  
Vol 81 (22) ◽  
pp. 7753-7766 ◽  
Author(s):  
Qian Liu ◽  
Yong Liang ◽  
Yun Zhang ◽  
Xiuling Shang ◽  
Shuwen Liu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Fluorescent Protein ◽  
Branched Chain Amino Acids ◽  
Transport Systems ◽  
Content Type ◽  
Overexpression Strain ◽  
Branched Chain ◽  
Amino Acid Efflux

ABSTRACTAmino acid efflux transport systems have important physiological functions and play vital roles in the fermentative production of amino acids. However, no methionine exporter has yet been identified inEscherichia coli. In this study, we identified a novel amino acid exporter, YjeH, inE. coli. TheyjeHoverexpression strain exhibited high tolerance to the structural analogues ofl-methionine and branched-chain amino acids, decreased intracellular amino acid levels, and enhanced export rates in the presence of a Met-Met, Leu-Leu, Ile-Ile, or Val-Val dipeptide, suggesting that YjeH functions as an exporter ofl-methionine and the three branched-chain amino acids. The export of the four amino acids in theyjeHoverexpression strain was competitively inhibited in relation to each other. The expression ofyjeHwas strongly induced by increasing cytoplasmic concentrations of substrate amino acids. Green fluorescent protein (GFP)-tagged YjeH was visualized by total internal reflection fluorescence microscopy to confirm the plasma membrane localization of YjeH. Phylogenetic analysis of transporters indicated that YjeH belongs to the amino acid efflux family of the amino acid/polyamine/organocation (APC) superfamily. Structural modeling revealed that YjeH has the typical “5 + 5” transmembrane α-helical segment (TMS) inverted-repeat fold of APC superfamily transporters, and its binding sites are strictly conserved. The enhanced capacity ofl-methionine export by the overexpression ofyjeHin anl-methionine-producing strain resulted in a 70% improvement in titer. This study supplements the transporter classification and provides a substantial basis for the application of the methionine exporter in metabolic engineering.

scholarly journals Basal-Level Effects of (p)ppGpp in the Absence of Branched-Chain Amino Acids in Actinobacillus pleuropneumoniae

2020 ◽  
Vol 202 (8) ◽  
Author(s):  
Gang Li ◽  
Qian Zhao ◽  
Tian Luan ◽  
Yangbo Hu ◽  
Yueling Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
De Novo ◽  
Model Organism ◽  
Stringent Response ◽  
Actinobacillus Pleuropneumoniae ◽  
Branched Chain Amino Acids ◽  
Content Type ◽  
Phenotypic Differences ◽  
Branched Chain

ABSTRACT The (p)ppGpp-mediated stringent response (SR) is a highly conserved regulatory mechanism in bacterial pathogens, enabling adaptation to adverse environments, and is linked to pathogenesis. Actinobacillus pleuropneumoniae can cause damage to the lungs of pigs, its only known natural host. Pig lungs are known to have a low concentration of free branched-chain amino acids (BCAAs) compared to the level in plasma. We had investigated the role for (p)ppGpp in viability and biofilm formation of A. pleuropneumoniae. Now, we sought to determine whether (p)ppGpp was a trigger signal for the SR in A. pleuropneumoniae in the absence of BCAAs. Combining transcriptome and phenotypic analyses of the wild type (WT) and an relA spoT double mutant [which does not produce (p)ppGpp], we found that (p)ppGpp could repress de novo purine biosynthesis and activate antioxidant pathways. There was a positive correlation between GTP and endogenous hydrogen peroxide content. Furthermore, the growth, viability, morphology, and virulence were altered by the inability to produce (p)ppGpp. Genes involved in the biosynthesis of BCAAs were constitutively upregulated, regardless of the existence of BCAAs, without accumulation of (p)ppGpp beyond a basal level. Collectively, our study shows that the absence of BCAAs was not a sufficient signal to trigger the SR in A. pleuropneumoniae. (p)ppGpp-mediated regulation in A. pleuropneumoniae is different from that described for the model organism Escherichia coli. Further work will establish whether the (p)ppGpp-dependent SR mechanism in A. pleuropneumoniae is conserved among other veterinary pathogens, especially those in the Pasteurellaceae family. IMPORTANCE (p)ppGpp is a key player in reprogramming transcriptomes to respond to nutritional challenges. Here, we present transcriptional and phenotypic differences of A. pleuropneumoniae grown in different chemically defined media in the absence of (p)ppGpp. We show that the deprivation of branched-chain amino acids (BCAAs) does not elicit a change in the basal-level (p)ppGpp, but this level is sufficient to regulate the expression of BCAA biosynthesis. The mechanism found in A. pleuropneumoniae is different from that of the model organism Escherichia coli but similar to that found in some Gram-positive bacteria. This study not only broadens the research scope of (p)ppGpp but also further validates the complexity and multiplicity of (p)ppGpp regulation in microorganisms that occupy different biological niches.

scholarly journals The ilvGMEDA Operon Is Regulated by Transcription Attenuation in Vibrio alginolyticus ZJ-T

2019 ◽  
Vol 85 (19) ◽  
Author(s):  
Yiqin Deng ◽  
Xing Luo ◽  
Mei Xie ◽  
Philippe Bouloc ◽  
Chang Chen ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Branched Chain Amino Acids ◽  
Vibrio Alginolyticus ◽  
Gram Negative ◽  
Content Type ◽  
Wide Range ◽  
Transcription Attenuation ◽  
Branched Chain ◽  
Fish And Shellfish

ABSTRACT Bacteria synthesize amino acids according to their availability in the environment or, in the case of pathogens, within the host. We explored the regulation of the biosynthesis of branched-chain amino acids (BCAAs) (l-leucine, l-valine, and l-isoleucine) in Vibrio alginolyticus, a marine fish and shellfish pathogen and an emerging opportunistic human pathogen. In this species, the ilvGMEDA operon encodes the main pathway for biosynthesis of BCAAs. Its upstream regulatory region shows no sequence similarity to the corresponding region in Escherichia coli or other Enterobacteriaceae, and yet we show that this operon is regulated by transcription attenuation. The translation of a BCAA-rich peptide encoded upstream of the structural genes provides an adaptive response similar to the E. coli canonical model. This study of a nonmodel Gram-negative organism highlights the mechanistic conservation of transcription attenuation despite the absence of primary sequence conservation. IMPORTANCE This study analyzes the regulation of the biosynthesis of branched-chain amino acids (leucine, valine, and isoleucine) in Vibrio alginolyticus, a marine bacterium that is pathogenic to fish and humans. The results highlight the conservation of the main regulatory mechanism with that of the enterobacterium Escherichia coli, suggesting that such a mechanism appeared early during the evolution of Gram-negative bacteria, allowing adaptation to a wide range of environments.

scholarly journals Analysis of the LIV System of Campylobacter jejuni Reveals Alternative Roles for LivJ and LivK in Commensalism beyond Branched-Chain Amino Acid Transport

2011 ◽  
Vol 193 (22) ◽  
pp. 6233-6243 ◽  
Author(s):  
Deborah A. Ribardo ◽  
David R. Hendrixson
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Campylobacter Jejuni ◽  
Transport System ◽  
Amino Acid Transport ◽  
Branched Chain Amino Acids ◽  
Acid Transport ◽  
Content Type ◽  
Branched Chain Amino Acid ◽  
Branched Chain

Campylobacter jejuniis a leading cause of diarrheal disease in humans and an intestinal commensal in poultry and other agriculturally important animals. These zoonotic infections result in significant amounts ofC. jejunipresent in the food supply to contribute to disease in humans. We previously found that a transposon insertion inCjj81176_1038, encoding a homolog of theEscherichia coliLivJ periplasmic binding protein of the leucine, isoleucine, and valine (LIV) branched-chain amino acid transport system, reduced the commensal colonization capacity ofC. jejuni81-176 in chicks.Cjj81176_1038is the first gene of a six-gene locus that encodes homologous components of theE. coliLIV system. By analyzing mutants with in-frame deletions of individual genes or pairs of genes, we found that this system constitutes a LIV transport system inC. jejuniresponsible for a high level of leucine acquisition and, to a lesser extent, isoleucine and valine acquisition. Despite each LIV protein being required for branched-chain amino acid transport, only the LivJ and LivK periplasmic binding proteins were required for wild-type levels of commensal colonization of chicks. All LIV permease and ATPase components were dispensable forin vivogrowth. These results suggest that the biological functions of LivJ and LivK for colonization are more complex than previously hypothesized and extend beyond a role for binding and acquiring branched-chain amino acids during commensalism. In contrast to other studies indicating a requirement and utilization of other specific amino acids for colonization, acquisition of branched-chain amino acids does not appear to be a determinant forC. jejuniduring commensalism.

Removal of infused amino acids by splanchnic and leg tissues in humans

1986 ◽  
Vol 250 (4) ◽  
pp. E407-E413 ◽  
Author(s):  
R. A. Gelfand ◽  
M. G. Glickman ◽  
R. Jacob ◽  
R. S. Sherwin ◽  
R. A. DeFronzo
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Uptake ◽  
Branched Chain Amino Acids ◽  
Acid Uptake ◽  
Acid Infusion ◽  
Amino Acid Infusion ◽  
Significant Uptake ◽  
Branched Chain

To compare the contributions of splanchnic and skeletal muscle tissues to the disposal of intravenously administered amino acids, regional amino acid exchange was measured across the splanchnic bed and leg in 11 normal volunteers. Postabsorptively, net release of amino acids by leg (largely alanine and glutamine) was complemented by the net splanchnic uptake of amino acids. Amino acid infusion via peripheral vein (0.2 g X kg-1 X h-1) caused a doubling of plasma insulin and glucagon levels and a threefold rise in blood amino acid concentrations. Both splanchnic and leg tissues showed significant uptake of infused amino acids. Splanchnic tissues accounted for approximately 70% of the total body amino acid nitrogen disposal; splanchnic uptake was greatest for the glucogenic amino acids but also included significant quantities of branched-chain amino acids. In contrast, leg amino acid uptake was dominated by the branched-chain amino acids. Based on the measured leg balance, body skeletal muscle was estimated to remove approximately 25-30% of the total infused amino acid load and approximately 65-70% of the infused branched-chain amino acids. Amino acid infusion significantly stimulated both the leg efflux and the splanchnic uptake of glutamine (not contained in the infusate). We conclude that when amino acids are infused peripherally in normal humans, splanchnic viscera (liver and gut) are the major sites of amino acid disposal.

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