scholarly journals Regulation of CodY Activity through Modulation of Intracellular Branched-Chain Amino Acid Pools

2010 ◽  
Vol 192 (24) ◽  
pp. 6357-6368 ◽  
Author(s):  
Shaun R. Brinsmade ◽  
Roelco J. Kleijn ◽  
Uwe Sauer ◽  
Abraham L. Sonenshein
Keyword(s):  
Amino Acids ◽  
Bacterial Species ◽  
Regulatory Protein ◽  
Fold Increase ◽  
Acid Synthesis ◽  
Branched Chain Amino Acids ◽  
Nucleoside Triphosphate ◽  
Isoleucine Leucine ◽  
Branched Chain

ABSTRACT In several Gram-positive bacterial species, the global transcriptional regulatory protein CodY adjusts the expression of many metabolic genes, apparently in response to changes in the pools of specific metabolites, i.e., the branched-chain amino acids (BCAAs) isoleucine, leucine, and valine (ILV) and the nucleoside triphosphate GTP. CodY not only responds to these metabolites as measured in vitro but also regulates the genes that direct their synthesis. We have constructed a set of strains lacking binding sites for the CodY protein in cis at loci coding for the ILV biosynthetic machinery, effectively overexpressing these genes in an attempt to modulate the ILV input signal to CodY. Metabolite analyses of strains derepressed for genes needed for ILV synthesis revealed more than a 6-fold increase in the valine pool and a 2-fold increase in the isoleucine and leucine pools. Accumulation of the branched-chain amino acids was accompanied by a 24-fold induction of the bkd operon (required for branched-chain fatty acid synthesis) and 6-fold hyperrepression of the CodY-regulated yhdG and yufN genes, demonstrating that CodY perceives intracellular fluctuations in at least one if its input signals. We conclude that changes in the rate of endogenous ILV synthesis serve as an important signal for CodY-mediated gene regulation.

scholarly journals Genetic and Biochemical Analysis of the Interaction of Bacillus subtilis CodY with Branched-Chain Amino Acids

2009 ◽  
Vol 191 (22) ◽  
pp. 6865-6876 ◽  
Author(s):  
Anuradha C. Villapakkam ◽  
Luke D. Handke ◽  
Boris R. Belitsky ◽  
Vladimir M. Levdikov ◽  
Anthony J. Wilkinson ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Branched Chain Amino Acids ◽  
Site Directed Mutagenesis ◽  
Binding Studies ◽  
Hydrophobic Pocket ◽  
Isoleucine Leucine ◽  
Branched Chain

ABSTRACT Bacillus subtilis CodY protein is a DNA-binding global transcriptional regulator that responds to branched-chain amino acids (isoleucine, leucine, and valine) and GTP. Crystal structure studies have shown that the N-terminal region of the protein includes a GAF domain that contains a hydrophobic pocket within which isoleucine and valine bind. This region is well conserved in CodY homologs. Site-directed mutagenesis was employed to understand the roles of some of the residues in the GAF domain and hydrophobic pocket in interaction with isoleucine and GTP. The F40A, F71E, and F98A forms of CodY were inactive in vivo. They were activatable by GTP but to a much lesser extent by branched-chain amino acids in vitro. The CodY mutant R61A retained partial repression of target promoters in vivo and was able to respond to GTP in vitro but also responded poorly to branched-chain amino acids in vitro unless GTP was simultaneously present. Thus, the GAF domain includes residues essential for full activation of CodY by branched-chain amino acids, but these residues are not critical for activation by GTP. Binding studies with branched-chain amino acids and their analogs revealed that an amino group at position 2 and a methyl group at position 3 of valine are critical components of the recognition of the amino acids by CodY.

Branched chain amino Acids as in vitro and in vivo Anti-Oxidation Compounds

2021 ◽  
pp. 3899-3904
Author(s):  
Moath Alqaraleh ◽  
Violet Kasabri ◽  
Ibrahim Al-Majali ◽  
Nihad Al-Othman ◽  
Nihad Al-Othman ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Amino Acids ◽  
Branched Chain Amino Acids ◽  
Raw 264.7 ◽  
Raw 264.7 Cell ◽  
Branched Chain ◽  
Raw 264.7 Cell Line

Background and aims: Branched chain amino acids (BCAAs) can be tightly connected to metabolism syndrome (MetS) which can be counted as a metabolic indicator in the case of insulin resistance (IR). The aim of this study was to assess the potential role of these acids under oxidative stress. Material and Methods: the in vitro antioxidant activity of BCAAs was assessed using free radical 1, 1-diphenyl-2-picryl-hydrazyl (DPPH) scavenging assays. For further check, a qRT-PCR technique was madefor detection the extent of alterations in gene expression of antioxidative enzymes (catalase and glutathione peroxidase (Gpx)) in lipopolysaccharides (LPS(-induced macrophages RAW 264.7 cell line. Additionally, BCAAs antioxidant activity was evaluated based on plasma H2O2 levels and xanthine oxidase (XO) activity in prooxidative LPS-treated mice. Results: Different concentrations of BCAAs affected on DPPH radical scavenging activity but to lesser extent than the ascorbic acid. Besides, BCAAs obviously upregulated the gene expression levels of catalases and Gpx in LPS-modulated macrophage RAW 264.7 cell line. In vivo BCAAs significantly minimized the level of plasma H2O2 as well as the activity of XO activity under oxidative stress. Conclusion: our current findings suggest that BCAAs supplementation may potentially serve as a therapeutic target for treatment of oxidative stress occurs with atherosclerosis, IR-diabetes, MetS and tumorigenesis.

A1151 EFFECT OF BRANCHED CHAIN AMINO ACIDS (BCAA) ON DIAPHRAGM FATIGUE IN VITRO

1990 ◽  
Vol 73 (3A) ◽  
pp. NA-NA
Author(s):  
H. Yamada ◽  
Y. Ohta ◽  
I. Chaudhry ◽  
H. Nagashima ◽  
J. Askanazi ◽  
...  
Keyword(s):  
Amino Acids ◽  
Branched Chain Amino Acids ◽  
Branched Chain ◽  
Diaphragm Fatigue

Abstract 86: Catabolic Remodeling of Branched Chain Amino Acids in Heart Failure

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Haipeng Sun ◽  
Meiyi Zhou ◽  
Chen Gao ◽  
Kristine Olson ◽  
Ji-Youn Youn ◽  
...  
Keyword(s):  
Heart Failure ◽  
Amino Acids ◽  
Regulatory Protein ◽  
Pressure Overload ◽  
Branched Chain Amino Acids ◽  
Atp Production ◽  
Genetic Ablation ◽  
Catabolic Genes ◽  
Metabolic Remodeling ◽  
Branched Chain

Metabolic remodeling is an integral part of heart failure. Although glucose and fatty acids metabolism have been extensively studied, little is known about the role of amino acids homeostasis in heart physiology and pathology. Branched chain amino acids (BCAAs), including leucine, isoleucine, and valine, are essential amino acids for both protein synthesis and cellular signaling. Elevated levels of BCAAs have been linked with heart failure. However, the underlying regulatory mechanism and functional significance of abnormal BCAA catabolism in heart failure have not been established. We found that genes involved in BCAA catabolism, including a key regulatory protein PP2Cm, are significantly down-regulated at mRNA as well as protein level in pressure-overload induced failing heart in mice. Furthermore, the concentrations of BCAA catabolic products branched-chain keto acids (BCKAs) are also elevated in heart tissues of post TAC mice. Interestingly, the down-regulation of BCAA catabolic genes mimics a similar expression pattern observed in fetal heart, suggesting that decreased BCAA catabolic activity is part of the metabolic remodeling in pathologically stressed heart from an adult to a fetal-like state. Genetic ablation of PP2Cm in mouse leads to defect in BCAA catabolism and accumulation of BCAAs and BCKAs in cardiac tissue and serum. PP2Cm deficient mice had lower cardiac contractility and higher susceptibility to develop heart failure under pressure overload. In addition, BCKAs treatment to isolated mitochondria resulted in lower oxygen consumption rate and ATP production. PP2Cm deficiency as well as BCKAs treatment induced oxidative stress in cardiomyocyte and antioxidant treatment ameliorated the development of heart failure in PP2Cm deficient animals. Together, these data indicated that BCAA catabolic remodeling is likely an integrated component of metabolic remodeling during heart failure. More importantly, mis-regulation of BCAA catabolism in heart promoted heart failure progression, involving direct impact on mitochondrial function and redox homeostasis in cardiomyocytes.

Properties of Acetate Kinase Isozymes and a Branched-Chain Fatty Acid Kinase from a Spirochete

1982 ◽  
Vol 152 (1) ◽  
pp. 246-254
Author(s):  
Caroline S. Harwood ◽  
Ercole Canale-Parola
Keyword(s):  
Amino Acids ◽  
Fatty Acids ◽  
Molecular Weight ◽  
Fatty Acid ◽  
Branched Chain Amino Acids ◽  
Chain Fatty Acid ◽  
Acetate Kinase ◽  
Nucleoside Triphosphate ◽  
Branched Chain ◽  
Chain Fatty Acids

Spirochete MA-2, which is anaerobic, ferments glucose, forming acetate as a major product. The spirochete also ferments (but does not utilize as growth substrates) small amounts of l -leucine, l -isoleucine, and l -valine, forming the branched-chain fatty acids isovalerate, 2-methylbutyrate, and isobutyrate, respectively, as end products. Energy generated through the fermentation of these amino acids is utilized to prolong cell survival under conditions of growth substrate starvation. A branched-chain fatty acid kinase and two acetate kinase isozymes were resolved from spirochete MA-2 cell extracts. Kinase activity was followed by measuring the formation of acyl phosphate from fatty acid and ATP. The branched-chain fatty acid kinase was active with isobutyrate, 2-methylbutyrate, isovalerate, butyrate, valerate, or propionate as a substrate but not with acetate as a substrate. The acetate kinase isozymes were active with acetate and propionate as substrates but not with longer-chain fatty acids as substrates. The acetate kinase isozymes and the branched-chain fatty acid kinase differed in nucleoside triphosphate and cation specificities. Each acetate kinase isozyme had an apparent molecular weight of approximately 125,000, whereas the branched-chain fatty acid kinase had a molecular weight of approximately 76,000. These results show that spirochete MA-2 synthesizes a branched-chain fatty acid kinase specific for leucine, isoleucine, and valine fermentation. It is likely that a phosphate branched-chain amino acids is also synthesized by spirochete MA-2. Thus, in spirochete MA-2, physiological mechanisms have evolved which serve specifically to generate maintenance energy from branched-chain amino acids.

scholarly journals Molecular Mechanisms Underlying the Positive Stringent Response of the Bacillus subtilis ilv-leu Operon, Involved in the Biosynthesis of Branched-Chain Amino Acids

2008 ◽  
Vol 190 (18) ◽  
pp. 6134-6147 ◽  
Author(s):  
Shigeo Tojo ◽  
Takenori Satomura ◽  
Kanako Kumamoto ◽  
Kazutake Hirooka ◽  
Yasutaro Fujita
Keyword(s):  
Amino Acids ◽  
Bacillus Subtilis ◽  
Transcription Initiation ◽  
Molecular Mechanisms ◽  
Stringent Response ◽  
Branched Chain Amino Acids ◽  
Base Substitution ◽  
Branched Chain

ABSTRACT Branched-chain amino acids are the most abundant amino acids in proteins. The Bacillus subtilis ilv-leu operon is involved in the biosynthesis of branched-chain amino acids. This operon exhibits a RelA-dependent positive stringent response to amino acid starvation. We investigated this positive stringent response upon lysine starvation as well as decoyinine treatment. Deletion analysis involving various lacZ fusions revealed two molecular mechanisms underlying the positive stringent response of ilv-leu, i.e., CodY-dependent and -independent mechanisms. The former is most likely triggered by the decrease in the in vivo concentration of GTP upon lysine starvation, GTP being a corepressor of the CodY protein. So, the GTP decrease derepressed ilv-leu expression through detachment of the CodY protein from its cis elements upstream of the ilv-leu promoter. By means of base substitution and in vitro transcription analyses, the latter (CodY-independent) mechanism was found to comprise the modulation of the transcription initiation frequency, which likely depends on fluctuation of the in vivo RNA polymerase substrate concentrations after stringent treatment, and to involve at least the base species of adenine at the 5′ end of the ilv-leu transcript. As discussed, this mechanism is presumably distinct from that for B. subtilis rrn operons, which involves changes in the in vivo concentration of the initiating GTP.

scholarly journals Mechanism of De Novo Branched-Chain Amino Acid Synthesis as an Alternative Electron Sink in Hypoxic Aspergillus nidulans Cells

2010 ◽  
Vol 76 (5) ◽  
pp. 1507-1515 ◽  
Author(s):  
Motoyuki Shimizu ◽  
Tatsuya Fujii ◽  
Shunsuke Masuo ◽  
Naoki Takaya
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Aspergillus Nidulans ◽  
De Novo ◽  
Acid Synthesis ◽  
Branched Chain Amino Acids ◽  
Amino Acid Synthesis ◽  
Lactate Dehydrogenase Activity ◽  
Branched Chain Amino Acid ◽  
Branched Chain

ABSTRACT Although branched-chain amino acids are synthesized as building blocks of proteins, we found that the fungus Aspergillus nidulans excretes them into the culture medium under hypoxia. The transcription of predicted genes for synthesizing branched-chain amino acids was upregulated by hypoxia. A knockout strain of the gene encoding the large subunit of acetohydroxy acid synthase (AHAS), which catalyzes the initial reaction of the synthesis, required branched-chain amino acids for growth and excreted very little of them. Pyruvate, a substrate for AHAS, increased the amount of hypoxic excretion in the wild-type strain. These results indicated that the fungus responds to hypoxia by synthesizing branched-chain amino acids via a de novo mechanism. We also found that the small subunit of AHAS regulated hypoxic branched-chain amino acid production as well as cellular AHAS activity. The AHAS knockout resulted in higher ratios of NADH/NAD+ and NADPH/NADP+ under hypoxia, indicating that the branched-chain amino acid synthesis contributed to NAD+ and NADP+ regeneration. The production of branched-chain amino acids and the hypoxic induction of involved genes were partly repressed in the presence of glucose, where cells produced ethanol and lactate and increased levels of lactate dehydrogenase activity. These indicated that hypoxic branched-chain amino acid synthesis is a unique alternative mechanism that functions in the absence of glucose-to-ethanol/lactate fermentation and oxygen respiration.

Bootstrapped neural network models for analyzing the responses of broiler chicks to dietary protein and branched chain amino acids

2014 ◽  
Vol 94 (1) ◽  
pp. 79-85 ◽  
Author(s):  
A. Faridi ◽  
A. Golian ◽  
A. Heravi Mousavi ◽  
J. France
Keyword(s):  
Neural Network ◽  
Amino Acids ◽  
Dietary Protein ◽  
Average Daily Gain ◽  
Network Models ◽  
Branched Chain Amino Acids ◽  
Broiler Chicks ◽  
Neural Network Models ◽  
Isoleucine Leucine ◽  
Branched Chain

Faridi, A., Golian, A., Heravi Mousavi, A. and France, J. 2014. Bootstrapped neural network models for analyzing the responses of broiler chicks to dietary protein and branched chain amino acids. Can. J. Anim. Sci. 94: 79–85. Reliable prediction of avian responses to dietary nutrients is essential for planning, management, and optimization activities in poultry nutrition. In this study, two bootstrapped neural network (BNN) models, each containing 100 separated neural networks (SNN), were developed for predicting average daily gain (ADG) and feed efficiency (FE) of broiler chicks in response to intake of protein and branched chain amino acids (BCAA) in the starter period. Using a re-sampling method, 100 different batches of data were generated for both the ADG and FE sets. Starting with 270 data lines extracted from eight studies in the literature, SNN models were trained, tested, and validated with 136, 67, and 67 data lines, respectively. All 200 SNN models developed, along with their respective BNN ones, were subjected to optimization (to find the optimum dietary protein and BCAA levels that maximize ADG and FE). Statistical analysis indicated that based on R 2, the BNN models were more accurate in 76 and 56 cases (out of 100) compared with the SNN models developed for ADG and FE, respectively. Optimization of the BNN models showed protein, isoleucine, leucine, and valine requirements for maximum ADG were 231.80, 9.05, 14.03 and 10.90 g kg−1 of diet, respectively. Also, maximum FE was obtained when the diet contained 232.30, 9.07, 14.50, and 11.04 g kg−1 of protein, isoleucine, leucine, and valine, respectively. The results of this study suggest that in meta-analytic modelling, bootstrap re-sampling algorithms should be used to better analyze available data and thereby take full advantage of them. This issue is of importance in the animal sciences as producing reliable data is both expensive and time-consuming.

Sign in / Sign up

Export Citation Format

Share Document