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Chen Chen - One of the best experts on this subject based on the ideXlab platform.
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pyroPhosphate Fructose 6 Phosphate 1 phosphotransferase pfp1 regulates starch biosynthesis and seed development via heterotetramer formation in rice oryza sativa l
Plant Biotechnology Journal, 2020Co-Authors: Chen Chen, Bingshu He, Peng Zhang, Chen Yang, Zhengwu Fang, Yongli QiaoAbstract:PyroPhosphate-Fructose 6-Phosphate 1-phosphotransferase (PFP1) reversibly converts Fructose 6-Phosphate and pyroPhosphate to Fructose 1, 6-bisPhosphate and orthoPhosphate during glycolysis, and has diverse functions in plants. However, mechanisms underlying the regulation of starch metabolism by PFP1 remain elusive. This study addressed the function of PFP1 in rice floury endosperm and defective grain filling. Compared with the wild type, pfp1-3 exhibited remarkably low grain weight and starch content, significantly increased protein and lipid content, and altered starch physicochemical properties and changes in embryo development. Map-based cloning revealed that pfp1-3 is a novel allele and encodes the regulatory beta-subunit of PFP1 (PFP1beta). Measurement of nicotinamide adenine dinucleotide (NAD+) showed that mutation of PFP1beta markedly decreased its enzyme activity. PFP1beta and three of four putative catalytic alpha-subunits of PFP1, PFP1alpha1, PFP1alpha2, and PFP1alpha4, interacted with each other to form a heterotetramer. Additionally, PFP1beta, PFP1alpha1 and PFP1alpha2 also formed homodimers. Furthermore, transcriptome analysis revealed that mutation of PFP1beta significantly altered expression of many essential enzymes in starch biosynthesis pathways. Concentrations of multiple lipid and glycolytic intermediates and trehalose metabolites were elevated in pfp1-3 endosperm, indicating that PFP1 modulates endosperm metabolism, potentially through reversible adjustments to metabolic fluxes. Taken together, these findings provide new insights into seed endosperm development and starch biosynthesis and will help in the breeding of rice cultivars with higher grain yield and quality.
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pyroPhosphate Fructose 6 Phosphate 1 phosphotransferase pfp1 regulates starch biosynthesis and seed development via heterotetramer formation in rice oryza sativa l
Plant Biotechnology Journal, 2020Co-Authors: Chen Chen, Peng Zhang, Xingxun Liu, Yujie Liu, Hongyan Yao, Junliang Yin, Xin WeiAbstract:PyroPhosphate-Fructose 6-Phosphate 1-phosphotransferase (PFP1) reversibly converts Fructose 6-Phosphate and pyroPhosphate to Fructose 1, 6-bisPhosphate and orthoPhosphate during glycolysis, and has diverse functions in plants. However, mechanisms underlying the regulation of starch metabolism by PFP1 remain elusive. This study addressed the function of PFP1 in rice floury endosperm and defective grain filling. Compared with the wild type, pfp1-3 exhibited remarkably low grain weight and starch content, significantly increased protein and lipid content, and altered starch physicochemical properties and changes in embryo development. Map-based cloning revealed that pfp1-3 is a novel allele and encodes the regulatory β-subunit of PFP1 (PFP1β). Measurement of nicotinamide adenine dinucleotide (NAD+) showed that mutation of PFP1β markedly decreased its enzyme activity. PFP1β and three of four putative catalytic α-subunits of PFP1, PFP1α1, PFP1α2, and PFP1α4, interacted with each other to form a heterotetramer. Additionally, PFP1β, PFP1α1 and PFP1α2 also formed homodimers. Furthermore, transcriptome analysis revealed that mutation of PFP1β significantly altered expression of many essential enzymes in starch biosynthesis pathways. Concentrations of multiple lipid and glycolytic intermediates and trehalose metabolites were elevated in pfp1-3 endosperm, indicating that PFP1 modulates endosperm metabolism, potentially through reversible adjustments to metabolic fluxes. Taken together, these findings provide new insights into seed endosperm development and starch biosynthesis and will help in the breeding of rice cultivars with higher grain yield and quality.
Peng Zhang - One of the best experts on this subject based on the ideXlab platform.
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pyroPhosphate Fructose 6 Phosphate 1 phosphotransferase pfp1 regulates starch biosynthesis and seed development via heterotetramer formation in rice oryza sativa l
Plant Biotechnology Journal, 2020Co-Authors: Chen Chen, Bingshu He, Peng Zhang, Chen Yang, Zhengwu Fang, Yongli QiaoAbstract:PyroPhosphate-Fructose 6-Phosphate 1-phosphotransferase (PFP1) reversibly converts Fructose 6-Phosphate and pyroPhosphate to Fructose 1, 6-bisPhosphate and orthoPhosphate during glycolysis, and has diverse functions in plants. However, mechanisms underlying the regulation of starch metabolism by PFP1 remain elusive. This study addressed the function of PFP1 in rice floury endosperm and defective grain filling. Compared with the wild type, pfp1-3 exhibited remarkably low grain weight and starch content, significantly increased protein and lipid content, and altered starch physicochemical properties and changes in embryo development. Map-based cloning revealed that pfp1-3 is a novel allele and encodes the regulatory beta-subunit of PFP1 (PFP1beta). Measurement of nicotinamide adenine dinucleotide (NAD+) showed that mutation of PFP1beta markedly decreased its enzyme activity. PFP1beta and three of four putative catalytic alpha-subunits of PFP1, PFP1alpha1, PFP1alpha2, and PFP1alpha4, interacted with each other to form a heterotetramer. Additionally, PFP1beta, PFP1alpha1 and PFP1alpha2 also formed homodimers. Furthermore, transcriptome analysis revealed that mutation of PFP1beta significantly altered expression of many essential enzymes in starch biosynthesis pathways. Concentrations of multiple lipid and glycolytic intermediates and trehalose metabolites were elevated in pfp1-3 endosperm, indicating that PFP1 modulates endosperm metabolism, potentially through reversible adjustments to metabolic fluxes. Taken together, these findings provide new insights into seed endosperm development and starch biosynthesis and will help in the breeding of rice cultivars with higher grain yield and quality.
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pyroPhosphate Fructose 6 Phosphate 1 phosphotransferase pfp1 regulates starch biosynthesis and seed development via heterotetramer formation in rice oryza sativa l
Plant Biotechnology Journal, 2020Co-Authors: Chen Chen, Peng Zhang, Xingxun Liu, Yujie Liu, Hongyan Yao, Junliang Yin, Xin WeiAbstract:PyroPhosphate-Fructose 6-Phosphate 1-phosphotransferase (PFP1) reversibly converts Fructose 6-Phosphate and pyroPhosphate to Fructose 1, 6-bisPhosphate and orthoPhosphate during glycolysis, and has diverse functions in plants. However, mechanisms underlying the regulation of starch metabolism by PFP1 remain elusive. This study addressed the function of PFP1 in rice floury endosperm and defective grain filling. Compared with the wild type, pfp1-3 exhibited remarkably low grain weight and starch content, significantly increased protein and lipid content, and altered starch physicochemical properties and changes in embryo development. Map-based cloning revealed that pfp1-3 is a novel allele and encodes the regulatory β-subunit of PFP1 (PFP1β). Measurement of nicotinamide adenine dinucleotide (NAD+) showed that mutation of PFP1β markedly decreased its enzyme activity. PFP1β and three of four putative catalytic α-subunits of PFP1, PFP1α1, PFP1α2, and PFP1α4, interacted with each other to form a heterotetramer. Additionally, PFP1β, PFP1α1 and PFP1α2 also formed homodimers. Furthermore, transcriptome analysis revealed that mutation of PFP1β significantly altered expression of many essential enzymes in starch biosynthesis pathways. Concentrations of multiple lipid and glycolytic intermediates and trehalose metabolites were elevated in pfp1-3 endosperm, indicating that PFP1 modulates endosperm metabolism, potentially through reversible adjustments to metabolic fluxes. Taken together, these findings provide new insights into seed endosperm development and starch biosynthesis and will help in the breeding of rice cultivars with higher grain yield and quality.
Georg A Sprenger - One of the best experts on this subject based on the ideXlab platform.
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acid base catalyst discriminates between a Fructose 6 Phosphate aldolase and a transaldolase
Chemcatchem, 2015Co-Authors: L Stellmacher, Tatjana Sandalova, Sebastian Leptihn, Gunter Schneider, Georg A Sprenger, Anne K. SamlandAbstract:The residues responsible for binding the catalytic water molecule were interchanged between the closely related enzymes Fructose 6-Phosphate aldolase A (FSAA) and transaldolase B (TalB) from Escherichia coli. In FSAA, this water molecule is bound by hydrogen bonds to the side chains of three residues (Gln59, Thr109 and Tyr131), whereas in TalB only two residues (Glu96 and Thr156) participate. Single and double variants were characterised with respect to Fructose 6-Phosphate aldolase and transaldolase activity, stability, pH dependence of activity, pKa value of the essential lysine residue and their three dimensional structure. The double variant TalBE96Q F178Y showed improved aldolase activity with an apparent kcat of 4.3 s−1. The experimentally determined pKa values of the catalytic lysine residue revealed considerable differences: In FSAA, this lysine residue is deprotonated at assay conditions (pKa 5.5) whereas it is protonated in TalB (pKa 9.3). Hence, a deprotonation of the catalytic lysine residue, which is a prerequisite for an efficient nucleophilic attack in TalB, is not necessary in FSAA. Based upon these results, we propose a new mechanism for FSAA with Tyr131 as general acid.
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replacement of a phenylalanine by a tyrosine in the active site confers Fructose 6 Phosphate aldolase activity to the transaldolase of escherichia coli and human origin
Journal of Biological Chemistry, 2008Co-Authors: Sarah Schneider, Tatjana Sandalova, Gunter Schneider, Georg A Sprenger, Anne K. SamlandAbstract:Abstract Based on a structure-assisted sequence alignment we designed 11 focused libraries at residues in the active site of transaldolase B from Escherichia coli and screened them for their ability to synthesize Fructose 6-Phosphate from dihydroxyacetone and glyceraldehyde 3-Phosphate using a newly developed color assay. We found one positive variant exhibiting a replacement of Phe178 to Tyr. This mutant variant is able not only to transfer a dihydroxyacetone moiety from a ketose donor, Fructose 6-Phosphate, onto an aldehyde acceptor, erythrose 4-Phosphate (14 units/mg), but to use it as a substrate directly in an aldolase reaction (7 units/mg). With a single amino acid replacement the Fructose-6-Phosphate aldolase activity was increased considerably (>70-fold compared with wild-type). Structural studies of the wild-type and mutant protein suggest that this is due to a different H-bond pattern in the active site leading to a destabilization of the Schiff base intermediate. Furthermore, we show that a homologous replacement has a similar effect in the human transaldolase Taldo1 (aldolase activity, 14 units/mg). We also demonstrate that both enzymes TalB and Taldo1 are recognized by the same polyclonal antibody.
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Replacement of a phenylalanine by a tyrosine in the active site confers Fructose-6-Phosphate aldolase activity to the transaldolase of Escherichia coli and human origin.
The Journal of biological chemistry, 2008Co-Authors: Sarah Schneider, Tatjana Sandalova, Gunter Schneider, Georg A Sprenger, Anne K. SamlandAbstract:Based on a structure-assisted sequence alignment we designed 11 focused libraries at residues in the active site of transaldolase B from Escherichia coli and screened them for their ability to synthesize Fructose 6-Phosphate from dihydroxyacetone and glyceraldehyde 3-Phosphate using a newly developed color assay. We found one positive variant exhibiting a replacement of Phe(178) to Tyr. This mutant variant is able not only to transfer a dihydroxyacetone moiety from a ketose donor, Fructose 6-Phosphate, onto an aldehyde acceptor, erythrose 4-Phosphate (14 units/mg), but to use it as a substrate directly in an aldolase reaction (7 units/mg). With a single amino acid replacement the Fructose-6-Phosphate aldolase activity was increased considerably (>70-fold compared with wild-type). Structural studies of the wild-type and mutant protein suggest that this is due to a different H-bond pattern in the active site leading to a destabilization of the Schiff base intermediate. Furthermore, we show that a homologous replacement has a similar effect in the human transaldolase Taldo1 (aldolase activity, 14 units/mg). We also demonstrate that both enzymes TalB and Taldo1 are recognized by the same polyclonal antibody.
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Fructose 6 Phosphate aldolase in organic synthesis preparation of d fagomine n alkylated derivatives and preliminary biological assays
Organic Letters, 2006Co-Authors: José A. Castillo, Georg A Sprenger, Teodor Parella, Jesús Joglar, Jordi Calveras, Josefina Casas, Montserrat Mitjans, Pilar M Vinardell, Tomoyuki Inoue, Pere ClapésAbstract:d-Fructose-6-Phosphate aldolase (FSA) mediates a novel straightforward two-step chemo-enzymatic synthesis of d-fagomine and some of its N-alkylated derivatives in 51% isolated yield and 99% de. The key step is the FSA-catalyzed aldol addition of simple dihydroxyacetone (DHA) to N-Cbz-3-aminopropanal. The use of FSA greatly simplifies the enzymatic procedures that used dihydroxyacetonePhosphate or DHA/esters. Some N-alkyl derivatives synthesized elicited antifungal and antibacterial activity as well as enhanced inhibitory activity, and selectivity against β-galactosidase and α-glucosidase.
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Fructose 6 Phosphate aldolase and 1 deoxy d xylulose 5 Phosphate synthase from escherichia coli as tools in enzymatic synthesis of 1 deoxysugars
Journal of Molecular Catalysis B-enzymatic, 2002Co-Authors: Martin Schurmann, Melanie Schürmann, Georg A SprengerAbstract:Abstract We cloned the genes for a novel Fructose 6-Phosphate aldolase (FSA) and for 1-deoxy- d -xylulose 5-Phosphate synthase (DXS) from Escherichia coli and investigated in their potential for enzymatic synthesis. FSA is the first example of a novel type of class I aldolases as it catalyzes the reversible formation of Fructose 6-Phosphate from dihydroxyacetone and d -glyceraldehyde 3-Phosphate. It utilizes several aldehydes as acceptor compounds, and interestingly, hydroxyacetone is an alternative donor which can be used to generate 1-deoxysugars. DXS catalyzes the decarboxylation of pyruvate and transfers the covalently bound thiamin diPhosphate-intermediate C 2 -moiety to d -glyceraldehyde 3-Phosphate. The reaction product, 1-deoxy- d -xylulose 5-Phosphate, is a precursor to isoprenoids and vitamins. DXS also uses other sugar Phosphates as well as short aldehydes as acceptor substrates. Apart from pyruvate, the two α-ketoacids hydroxypyruvate and α-oxobutyrate could be used as donor substrates. FSA and DXS were successfully used to synthesize 1-deoxyketoses from C 4 (1-deoxy-erythrulose) to C 7 (1-deoxy-sedoheptulose) in phosphorylated and non-phosphorylated form.
Xin Wei - One of the best experts on this subject based on the ideXlab platform.
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pyroPhosphate Fructose 6 Phosphate 1 phosphotransferase pfp1 regulates starch biosynthesis and seed development via heterotetramer formation in rice oryza sativa l
Plant Biotechnology Journal, 2020Co-Authors: Chen Chen, Peng Zhang, Xingxun Liu, Yujie Liu, Hongyan Yao, Junliang Yin, Xin WeiAbstract:PyroPhosphate-Fructose 6-Phosphate 1-phosphotransferase (PFP1) reversibly converts Fructose 6-Phosphate and pyroPhosphate to Fructose 1, 6-bisPhosphate and orthoPhosphate during glycolysis, and has diverse functions in plants. However, mechanisms underlying the regulation of starch metabolism by PFP1 remain elusive. This study addressed the function of PFP1 in rice floury endosperm and defective grain filling. Compared with the wild type, pfp1-3 exhibited remarkably low grain weight and starch content, significantly increased protein and lipid content, and altered starch physicochemical properties and changes in embryo development. Map-based cloning revealed that pfp1-3 is a novel allele and encodes the regulatory β-subunit of PFP1 (PFP1β). Measurement of nicotinamide adenine dinucleotide (NAD+) showed that mutation of PFP1β markedly decreased its enzyme activity. PFP1β and three of four putative catalytic α-subunits of PFP1, PFP1α1, PFP1α2, and PFP1α4, interacted with each other to form a heterotetramer. Additionally, PFP1β, PFP1α1 and PFP1α2 also formed homodimers. Furthermore, transcriptome analysis revealed that mutation of PFP1β significantly altered expression of many essential enzymes in starch biosynthesis pathways. Concentrations of multiple lipid and glycolytic intermediates and trehalose metabolites were elevated in pfp1-3 endosperm, indicating that PFP1 modulates endosperm metabolism, potentially through reversible adjustments to metabolic fluxes. Taken together, these findings provide new insights into seed endosperm development and starch biosynthesis and will help in the breeding of rice cultivars with higher grain yield and quality.
Kerry S. Smith - One of the best experts on this subject based on the ideXlab platform.
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allosteric regulation of lactobacillus plantarum xylulose 5 Phosphate Fructose 6 Phosphate phosphoketolase xfp
Journal of Bacteriology, 2015Co-Authors: Katie Glenn, Kerry S. SmithAbstract:ABSTRACT Xylulose 5-Phosphate/Fructose 6-Phosphate phosphoketolase (Xfp), which catalyzes the conversion of xylulose 5-Phosphate (X5P) or Fructose 6-Phosphate (F6P) to acetyl Phosphate, plays a key role in carbohydrate metabolism in a number of bacteria. Recently, we demonstrated that the fungal Cryptococcus neoformans Xfp2 exhibits both substrate cooperativity for all substrates (X5P, F6P, and P i ) and allosteric regulation in the forms of inhibition by phosphoenolpyruvate (PEP), oxaloacetic acid (OAA), and ATP and activation by AMP (K. Glenn, C. Ingram-Smith, and K. S. Smith. Eukaryot Cell 13: 657–663, 2014). Allosteric regulation has not been reported previously for the characterized bacterial Xfps. Here, we report the discovery of substrate cooperativity and allosteric regulation among bacterial Xfps, specifically the Lactobacillus plantarum Xfp. L. plantarum Xfp is an allosteric enzyme inhibited by PEP, OAA, and glyoxylate but unaffected by the presence of ATP or AMP. Glyoxylate is an additional inhibitor to those previously reported for C. neoformans Xfp2. As with C. neoformans Xfp2, PEP and OAA share the same or possess overlapping sites on L. plantarum Xfp. Glyoxylate, which had the lowest half-maximal inhibitory concentration of the three inhibitors, binds at a separate site. This study demonstrates that substrate cooperativity and allosteric regulation may be common properties among bacterial and eukaryotic Xfp enzymes, yet important differences exist between the enzymes in these two domains. IMPORTANCE Xylulose 5-Phosphate/Fructose 6-Phosphate phosphoketolase (Xfp) plays a key role in carbohydrate metabolism in a number of bacteria. Although we recently demonstrated that the fungal Cryptococcus Xfp is subject to substrate cooperativity and allosteric regulation, neither phenomenon has been reported for a bacterial Xfp. Here, we report that the Lactobacillus plantarum Xfp displays substrate cooperativity and is allosterically inhibited by phosphoenolpyruvate and oxaloacetate, as is the case for Cryptococcus Xfp. The bacterial enzyme is unaffected by the presence of AMP or ATP, which act as a potent activator and inhibitor of the fungal Xfp, respectively. Our results demonstrate that substrate cooperativity and allosteric regulation may be common properties among bacterial and eukaryotic Xfps, yet important differences exist between the enzymes in these two domains.
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biochemical and kinetic characterization of xylulose 5 Phosphate Fructose 6 Phosphate phosphoketolase 2 xfp2 from cryptococcus neoformans
Eukaryotic Cell, 2014Co-Authors: Katie Glenn, Cheryl Ingramsmith, Kerry S. SmithAbstract:Xylulose 5-Phosphate/Fructose 6-Phosphate phosphoketolase (Xfp), previously thought to be present only in bacteria but recently found in fungi, catalyzes the formation of acetyl Phosphate from xylulose 5-Phosphate or Fructose 6-Phosphate. Here, we describe the first biochemical and kinetic characterization of a eukaryotic Xfp, from the opportunistic fungal pathogen Cryptococcus neoformans, which has two XFP genes (designated XFP1 and XFP2). Our kinetic characterization of C. neoformans Xfp2 indicated the existence of both substrate cooperativity for all three substrates and allosteric regulation through the binding of effector molecules at sites separate from the active site. Prior to this study, Xfp enzymes from two bacterial genera had been characterized and were determined to follow Michaelis-Menten kinetics. C. neoformans Xfp2 is inhibited by ATP, phosphoenolpyruvate (PEP), and oxaloacetic acid (OAA) and activated by AMP. ATP is the strongest inhibitor, with a half-maximal inhibitory concentration (IC50) of 0.6 mM. PEP and OAA were found to share the same or have overlapping allosteric binding sites, while ATP binds at a separate site. AMP acts as a very potent activator; as little as 20 μM AMP is capable of increasing Xfp2 activity by 24.8% ± 1.0% (mean ± standard error of the mean), while 50 μM prevented inhibition caused by 0.6 mM ATP. AMP and PEP/OAA operated independently, with AMP activating Xfp2 and PEP/OAA inhibiting the activated enzyme. This study provides valuable insight into the metabolic role of Xfp within fungi, specifically the fungal pathogen Cryptococcus neoformans, and suggests that at least some Xfps display substrate cooperative binding and allosteric regulation.
