- Chemical and enzymatic methodologies for the synthesis of enantiomerically pure glyceraldehyde 3-phosphates
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Glyceraldehyde 3-phosphates are important intermediates of many central metabolic pathways in a large number of living organisms. d-Glyceraldehyde 3-phosphate (d-GAP) is a key intermediate during glycolysis and can as well be found in a variety of other metabolic pathways. The opposite enantiomer, l-glyceraldehyde 3-phosphate (l-GAP), has been found in a few exciting new pathways. Here, improved syntheses of enantiomerically pure glyceraldehyde 3-phosphates are reported. While d-GAP was synthesized by periodate cleavage of d-fructose 6-phosphate, l-GAP was obtained by enzymatic phosphorylation of l-glyceraldehyde. 1H- and 31P NMR spectroscopy was applied in order to examine pH-dependent behavior of GAP over time and to identify potential degradation products. It was found that GAP is stable in acidic aqueous solution below pH 4. At pH 7, methylglyoxal is formed, whereas under alkaline conditions, the formation of lactic acid could be observed.
- Gauss, Dominik,Schoenenberger, Bernhard,Wohlgemuth, Roland
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- Rational engineering of 2-deoxyribose-5-phosphate aldolases for the biosynthesis of (R)-1,3-butanediol
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Carbon– carbon bond formation is one of the most important reactions in biocatalysis and organic chemistry. In nature, aldolases catalyze the reversible stereoselective aldol addition between two carbonyl compounds, making them attractive catalysts for the synthesis of various chemicals. In this work, we identified several 2-deoxyribose-5-phosphate aldolases (DERAs) having acetaldehyde condensation activity, which can be used for the biosynthesis of (R)-1,3-butanediol (1,3BDO) in combination with aldo-keto reductases (AKRs). Enzymatic screening of 20 purified DERAs revealed the presence of significant acetaldehyde condensation activity in 12 of the enzymes, with the highest activities in BH1352 from Bacillus halodurans, TM1559 from Thermotoga maritima, and DeoC from Escherichia coli. The crystal structures of BH1352 and TM1559 at 1.40 –2.50 ? resolution are the first full-length DERA structures revealing the presence of the C-terminal Tyr (Tyr224 in BH1352). The results from structure-based site-directed mutagenesis of BH1352 indicated a key role for the catalytic Lys155 and other active-site residues in the 2-deoxyribose-5-phosphate cleavage and acetaldehyde condensation reactions. These experiments also revealed a 2.5-fold increase in acetaldehyde transformation to 1,3BDO (in combination with AKR) in the BH1352 F160Y and F160Y/M173I variants. The replacement of the WT BH1352 by the F160Y or F160Y/M173I variants in E. coli cells expressing the DERA + AKR pathway increased the production of 1,3BDO from glucose five and six times, respectively. Thus, our work provides detailed insights into the molecular mechanisms of substrate selectivity and activity of DERAs and identifies two DERA variants with enhanced activity for in vitro and in vivo 1,3BDO biosynthesis.
- Kim, Taeho,Stogios, Peter J.,Khusnutdinova, Anna N.,Nemr, Kayla,Skarina, Tatiana,Flick, Robert,Joo, Jeong Chan,Mahadevan, Radhakrishnan,Savchenko, Alexei,Yakunin, Alexander F.
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- N-sulfonyl hydroxamate derivatives as inhibitors of class II fructose-1,6-diphosphate aldolase
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Dihydroxyacetone-phosphate and phosphonate derivatives were synthesized bearing a N-sulfonyl hydroxamate moiety. The phosphate derivatives represent competitive inhibitors for the class II-FBP aldolase catalyzed reaction, while the phosphonate isosteres are comparatively weaker inhibitors.
- Gavalda, Sabine,Braga, Remi,Dax, Chantal,Vigroux, Alain,Blonski, Casimir
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- Substitutions at a rheostat position in human aldolase A cause a shift in the conformational population
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Some protein positions play special roles in determining the magnitude of protein function: at such “rheostat” positions, varied amino acid substitutions give rise to a continuum of functional outcomes, from wild type (or enhanced), to intermediate, to loss of function. This observed range raises interesting questions about the biophysical bases by which changes at single positions have such varied outcomes. Here, we assessed variants at position 98 in human aldolase A (“I98X”). Despite being ~17 ? from the active site and far from subunit interfaces, substitutions at position 98 have rheostatic contributions to the apparent cooperativity (nH) associated with fructose-1,6-bisphosphate substrate binding and moderately affected binding affinity. Next, we crystallized representative I98X variants to assess structural consequences. Residues smaller than the native isoleucine (cysteine and serine) were readily accommodated, and the larger phenylalanine caused only a slight separation of the two parallel helixes. However, the diffraction quality was reduced for I98F, and further reduced for I98Y. Intriguingly, the resolutions of the I98X structures correlated with their nH values. We propose that substitution effects on both nH and crystal lattice disruption arise from changes in the population of aldolase A conformations in solution. In combination with results computed for rheostat positions in other proteins, the results from this study suggest that rheostat positions accommodate a wide range of side chains and that structural consequences manifest as shifted ensemble populations and/or dynamics changes.
- Fenton, Aron W.,Fenton, Kathryn D.,Lamb, Audrey L.,Martin, Tyler A.,Meneely, Kathleen M.,Swint-Kruse, Liskin,Wu, Tiffany
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- Prebiotic synthesis of aminooxazoline-5′-phosphates in water by oxidative phosphorylation
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RNA is essential to all life on Earth and is the leading candidate for the first biopolymer of life. Aminooxazolines have recently emerged as key prebiotic ribonucleotide precursors, and here we develop a novel strategy for aminooxazoline-5′-phosphate synthesis in water from prebiotic feedstocks. Oxidation of acrolein delivers glycidaldehyde (90%), which directs a regioselective phosphorylation in water and specifically affords 5′-phosphorylated nucleotide precursors in upto 36% yield. We also demonstrated a generational link between proteinogenic amino acids (Met, Glu, Gln) and nucleotide synthesis.
- Fernández-García,Grefenstette,Powner
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supporting information
p. 4919 - 4921
(2017/07/11)
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- One-pot enzymatic reaction sequence for the syntheses of d-glyceraldehyde 3-phosphate and l-glycerol 3-phosphate
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A one-pot enzymatic reaction sequence for the synthesis of optically pure d-glyceraldehyde 3-phosphate (d-GAP) and l-glycerol 3-phosphate (sn-G3P) was designed using fructose-bisphosphate aldolase from rabbit muscle (RAMA), sn-glycerol 3-phosphate dehydrogenase (sn-G3PDH) and formate dehydrogenase from Candida boidinii (FDH). The reaction sequence significantly improves the aldol cleavage of d-fructose 1,6-bisphosphate (d-F16BP) catalyzed by RAMA and yields 100% conversion of d-F16BP by overcoming thermodynamic limitation. The degradation kinetics of d-GAP under reaction conditions was investigated and a reaction kinetics model defining the entire cascade was developed. Validation of the model shows 98.5% correlation between experimental data and numerically simulated data matrices. The evaluation of different types of reactor was performed by combining the reaction kinetics model, mass balances and kinetics of the non-enzymatic degradation of d-GAP. Batch-wise operation in a stirred tank reactor (STR) is the most convenient procedure for the one-pot enzymatic syntheses of d-GAP and sn-G3P. The separation of the two products d-GAP and sn-G3P has been achieved using polyethylenimine (PEI)-cellulose TLC.
- Molla, Getachew S.,Wohlgemuth, Roland,Liese, Andreas
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- Dual Activity of Quinolinate Synthase: Triose Phosphate Isomerase and Dehydration Activities Play Together to Form Quinolinate
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Quinolinate synthase (NadA) is an Fe4S4 cluster-containing dehydrating enzyme involved in the synthesis of quinolinic acid (QA), the universal precursor of the essential coenzyme nicotinamide adenine dinucleotide. The reaction catalyzed by NadA is not well understood, and two mechanisms have been proposed in the literature that differ in the nature of the molecule (DHAP or G-3P) that condenses with iminoaspartate (IA) to form QA. In this article, using biochemical approaches, we demonstrate that DHAP is the triose that condenses with IA to form QA. The capacity of NadA to use G-3P is due to its previously unknown triose phosphate isomerase activity.
- Reichmann, Debora,Couté, Yohann,Ollagnier De Choudens, Sandrine
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p. 6443 - 6446
(2015/11/09)
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- Structural mutations that probe the interactions between the catalytic and dianion activation sites of triosephosphate isomerase
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Triosephosphate isomerase (TIM) catalyzes the isomerization of dihydroxyacetone phosphate to form d-glyceraldehyde 3-phosphate. The effects of two structural mutations in TIM on the kinetic parameters for catalysis of the reaction of the truncated substrate glycolaldehyde (GA) and the activation of this reaction by phosphite dianion are reported. The P168A mutation results in similar 50- and 80-fold decreases in (kcat/Km)E and (kcat/Km)E·HPi, respectively, for deprotonation of GA catalyzed by free TIM and by the TIM·HPO 32- complex. The mutation has little effect on the observed and intrinsic phosphite dianion binding energy or the magnitude of phosphite dianion activation of TIM for catalysis of deprotonation of GA. A loop 7 replacement mutant (L7RM) of TIM from chicken muscle was prepared by substitution of the archaeal sequence 208-TGAG with 208-YGGS. L7RM exhibits a 25-fold decrease in (kcat/Km)E and a larger 170-fold decrease in (kcat/Km)E·HPi for reactions of GA. The mutation has little effect on the observed and intrinsic phosphodianion binding energy and only a modest effect on phosphite dianion activation of TIM. The observation that both the P168A and loop 7 replacement mutations affect mainly the kinetic parameters for TIM-catalyzed deprotonation but result in much smaller changes in the parameters for enzyme activation by phosphite dianion provides support for the conclusion that catalysis of proton transfer and dianion activation of TIM take place at separate, weakly interacting, sites in the protein catalyst.
- Zhai, Xiang,Amyes, Tina L.,Wierenga, Rik K.,Loria, J. Patrick,Richard, John P.
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p. 5928 - 5940
(2013/09/23)
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- Kinetic and mechanistic characterization of the glyceraldehyde 3-phosphate dehydrogenase from Mycobacterium tuberculosis
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Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is a glycolytic protein responsible for the conversion of glyceraldehyde 3-phosphate (G3P), inorganic phosphate and nicotinamide adenine dinucleotide (NAD+) to 1,3-bisphosphoglycerate (1,3-BPG) and the reduced form of nicotinamide adenine dinucleotide (NADH). Here we report the characterization of GAPDH from Mycobacterium tuberculosis (Mtb). This enzyme exhibits a kinetic mechanism in which first NAD+, then G3P bind to the active site resulting in the formation of a covalently bound thiohemiacetal intermediate. After oxidation of the thiohemiacetal and subsequent nucleotide exchange (NADH off, NAD+ on), the binding of inorganic phosphate and phosphorolysis yields the product 1,3-BPG. Mutagenesis and iodoacetamide (IAM) inactivation studies reveal the conserved C158 to be responsible for nucleophilic catalysis and that the conserved H185 to act as a catalytic base. Primary, solvent and multiple kinetic isotope effects revealed that the first half-reaction is rate limiting and utilizes a step-wise mechanism for thiohemiacetal oxidation via a transient alkoxide to promote hydride transfer and thioester formation.
- Wolfson-Stofko, Brett,Hadi, Timin,Blanchard, John S.
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- FSAB: A new fructose-6-phosphate aldolase from Escherichia coli. Cloning, over-expression and comparative kinetic characterization with FSAA
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Fructose-6-phosphate aldolase B (FSAB) from Escherichia coli was successfully over-expressed as His-tagged recombinant protein. A decameric protein was observed as for FSAA. Unlike FSAA, FSAB is not thermally stable at temperatures higher than 60 °C. The 70% identity between the two aldolases has allowed the generation of a 3D structure which has shown a high similarity of the two active sites. Full kinetic studies towards several substrates have revealed that FSAB catalytic activity is very close to FSAA activity, corroborated by the similarity of their active sites. FSAB has been able to react with three known donors (dihydroxyacetone, hydroxyacetone and glycolaldehyde) but always slightly slower than FSAA.
- Sanchez-Moreno, Israel,Nauton, Lionel,Thery, Vincent,Pinet, Agnes,Petit, Jean-Louis,De Berardinis, Veronique,Samland, Anne K.,Guerard-Helaine, Christine,Lemaire, Marielle
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- Wildtype and engineered monomeric triosephosphate isomerase from Trypanosoma brucei: Partitioning of reaction intermediates in D2O and activation by phosphite dianion
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Product yields for the reactions of (R)-glyceraldehyde 3-phosphate (GAP) in D2O at pD 7.9 catalyzed by wildtype triosephosphate isomerase from Trypanosoma brucei brucei (Tbb TIM) and a monomeric variant (monoTIM) of this wildtype enzyme were determined by 1H NMR spectroscopy and were compared with the yields determined in earlier work for the reactions catalyzed by TIM from rabbit and chicken muscle [O'Donoghue, A. C., Amyes, T. L., and Richard, J. P. (2005), Biochemistry44, 2610 -2621]. Three products were observed from the reactions catalyzed by TIM: dihydroxyacetone phosphate (DHAP) from isomerization with intramolecular transfer of hydrogen, d-DHAP from isomerization with incorporation of deuterium from D2O into C-1 of DHAP, and d-GAP from incorporation of deuterium from D2O into C-2 of GAP. The yield of DHAP formed by intramolecular transfer of hydrogen decreases from 49% for the muscle enzymes to 40% for wildtype Tbb TIM to 34% for monoTIM. There is no significant difference in the ratio of the yields of d-DHAP and d-GAP for wildtype TIM from muscle sources and Trypanosoma brucei brucei, but partitioning of the enediolate intermediate of the monoTIM reaction to form d-DHAP is less favorable ((kC1)D/(kC2) D = 1.1) than for the wildtype enzyme ((kC1) D/(kC2)D = 1.7). Product yields for the wildtype Tbb TIM and monoTIM-catalyzed reactions of glycolaldehyde labeled with carbon-13 at the carbonyl carbon ([1-13C]-GA) at pD 7.0 in the presence of phosphite dianion and in its absence were determined by 1H NMR spectroscopy [Go, M. K., Amyes, T. L., and Richard, J. P. (2009) Biochemistry48, 5769-5778]. There is no detectable difference in the yields of the products of wildtype muscle and Tbb TIM-catalyzed reactions of [1-13C]-GA in D2O. The kinetic parameters for phosphite dianion activation of the reactions of [1-13C]-GA catalyzed by wildtype Tbb TIM are similar to those reported for the enzyme from rabbit muscle [Amyes, T. L. and Richard, J. P. (2007) Biochemistry46, 5841-5854], but there is no detectable dianion activation of the reaction catalyzed by monoTIM. The engineered disruption of subunit contacts at monoTIM causes movement of the essential side chains of Lys-13 and His-95 away from the catalytic active positions. We suggest that this places an increased demand that the intrinsic binding energy of phosphite dianion be utilized to drive the change in the conformation of monoTIM back to the active structure for wildtype TIM.
- Malabanan, M. Merced,Go, Maybelle K.,Amyes, Tina L.,Richard, John P.
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p. 5767 - 5779
(2012/05/20)
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- An improved preparation of D-glyceraldehyde 3-phosphate and its use in the synthesis of 1-deoxy-D-xylulose 5-phosphate
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D-Glyceraldehyde 3-phosphate (=D-GAP; 2) was prepared by an improved chemical method (Scheme 2), and it was then employed to synthesize 1-deoxy-d-xylulose 5-phosphate (=DXP; 3) which is enzymatically one of the key intermediates in the MEP (4) terpenoid b
- Li, Heng,Tian, Jie,Wang, Hui,Yang, Shao-Qing,Gao, Wen-Yun
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experimental part
p. 1745 - 1750
(2010/12/20)
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- Cloning, expression, purification, cofactor requirements, and steady state kinetics of phosphoketolase-2 from Lactobacillus plantarum
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The genes xpk1 and xpk2(Δ1-21) encoding phosphoketolase-1 and (Δ1-7)-truncated phosphoketolase-2 have been cloned from Lactobacillus plantarum and expressed in Escherichia coli. Both gene-products display phosphoketolase activity on fructose-6-phosphate in extracts. A N-terminal His-tag construct of xpk2(Δ1-21) was also expressed in E. coli and produced active His-tagged (Δ1-7)-truncated phosphoketolase-2 (hereafter phosphoketolase-2). Phosphoketolase-2 is activated by thiamine pyrophosphate (TPP) and the divalent metal ions Mg2+, Mn2+, or Ca2+. Kinetic analysis and data from the literature indicate the activators are MgTPP, MnTPP, or CaTPP, and these species activate by an ordered equilibrium binding pathway, with Me2+TPP binding first and then fructose-6-phosphate. Phosphoketolase-2 accepts either fructose-6-phosphate or xylulose-5-phosphate as substrates, together with inorganic phosphate, to produce acetyl phosphate and either erythrose-4-phosphate or glyceraldehyde-3-phosphate, respectively. Steady state kinetic analysis of acetyl phosphate formation with either substrate indicates a ping pong kinetic mechanism. Product inhibition patterns with erythrose-4-phosphate indicate that an intermediate in the ping pong mechanism is formed irreversibly. Background mechanistic information indicates that this intermediate is 2-acetyl-TPP. The irreversibility of 2-acetyl-TPP formation might explain the overall irreversibility of the reaction of phosphoketolase-2.
- Yevenes, Alejandro,Frey, Perry A.
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p. 121 - 127
(2008/09/20)
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- Separate synthesis and evaluation of glucitol bis-phosphate and mannitol bis-phosphate, as competitive inhibitors of fructose bis-phosphate aldolases
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We report the first unambiguous syntheses of glucitol-1,6-bis-phosphate and mannitol-1,6-bis-phosphate and their competitive inhibition of various fructose bis-phosphate aldolases.
- Mabiala-Bassiloua, Charles-Gabin,Zwolinska, Magdalena,Therisod, Helene,Sygusch, Jurgen,Therisod, Michel
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p. 1735 - 1737
(2008/12/21)
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- Evolution of the indole alkaloid biosynthesis in the genus Hordeum: Distribution of gramine and DIBOA and isolation of the benzoxazinoid biosynthesis genes from Hordeum lechleri
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Two indole alkaloids with defense related functions are synthesized in the genus Hordeum of the Triticeae. Gramine (3(dimethyl-amino-methyl)-indole) is found in H. spontaneum and in some varieties of H. vulgare, the benzoxazinoid 2,4-dihydroxy-2H-1,4-benzoxazin-3(4H)-one (DIBOA) is detected in H. roshevitzii, H. brachyantherum, H. flexuosum, H. lechleri. Biosynthesis of DIBOA and of gramine was found to be mutually exclusive in wild Hordeum species, indicating that there was selection against simultaneous expression of both pathways during evolution. The full set of genes required for DIBOA biosynthesis in H.lechleri was isolated and the respective enzyme functions were analyzed by heterologous expression. The cytochrome P450 genes Bx2-Bx5 demonstrate a monophyletic origin for H. lechleri, Triticum aestivum and Zea mays. HlBx2-HlBx5 share highest homology to the orthologous genes of T. aestivum. In contrast, the branch point enzyme of the DIBOA pathway, the indole-3-glycerol phosphate lyase BX1, might have evolved independently in H. lechleri. In all Hordeum species that synthesize DIBOA, DNA sequences homologous to Bx genes are found. In contrast, these sequences are not detectable in the genomes of H. vulgare and H. spontaneum that do not synthesize benzoxazinoids.
- Gruen, Sebastian,Frey, Monika,Gierl, Alfons
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p. 1264 - 1272
(2007/10/03)
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- Purification and characterization of an allosteric fructose-1,6- bisphosphate aldolase from germinating mung beans (Vigna radiata)
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Cytosolic fructose-1,6-P2 (FBP) aldolase (ALDc) from germinated mung beans has been purified 1078-fold to electrophoretic homogeneity and a final specific activity of 15.1 μmol FBP cleaved/min per mg of protein. SDS-PAGE of the final preparation revealed a single protein-staining band of 40 kDa that cross-reacted strongly with rabbit anti-(carrot ALD c)-IgG. The enzyme's native Mr was determined by gel filtration chromatography to be 160 kDa, indicating a homotetrameric quaternary structure. This ALD is a class I ALD, since EDTA or Mg2+ had no effect on its activity, and was relatively heat-stable losing 0-25% of its activity when incubated for 5 min at 55-65°C. It demonstrated: (i) a temperature coefficient (Q10) of 1.7; (ii) an activation energy of 9.2 kcal/mol active site; and (iii) a broad pH-activity optima of 7.5. Mung bean ALDc is bifunctional for FBP and sedoheptulose-1,7-P2 (Km ≈ 17 μM for both substrates). ATP, ADP, AMP and ribose-5-P exerted inhibitory effects on the activity of the purified enzyme. Ribose-5-P, ADP and AMP functioned as competitive inhibitors (Ki values = 2.2, 3.1 and 7.5 mM, respectively). By contrast, the addition of 2 mM ATP: (i) reduced Vmax by about 2-fold, (ii) increased Km(FBP) by about 4-fold, and (iii) shifted the FBP saturation kinetic plot from hyperbolic to sigmoidal (h = 1.0 and 2.6 in the absence and presence of 2 mM ATP, respectively). Potent feedback inhibition of ALDc by ATP is suggested to help balance cellular ATP demands with the control of cytosolic glycolysis and respiration in germinating mung beans.
- Lal, Ashish,Plaxton, William C.,Kayastha, Arvind M.
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p. 968 - 974
(2007/10/03)
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- Slow-binding inhibition of 2-keto-3-deoxy-6-phosphogluconate (KDPG) aldolase
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2-Keto-3-deoxy-6-phosphogluconate (KDPG) aldolase is a key enzyme in the Entner-Doudoroff pathway of bacteria. It catalyzes the reversible production of KDPG from pyruvate and D-glyceraldehyde 3-phosphate through a class I Schiff base mechanism. On the basis of aldolase mechanistic pathway, various pyruvate analogues bearing β-diketo structures were designed and synthesized as potential inhibitors. Their capacity to inhibit aldolase catalyzed reaction by forming stabilized iminium ion or conjugated enamine were investigated by enzymatic kinetics and UV-vis difference spectroscopy. Depending of the substituent R (methyl or aromatic ring), a competitive or a slow-binding inhibition takes place. These results were examined on the basis of the three-dimensional structure of the enzyme.
- Braga, Remi,Hecquet, Laurence,Blonski, Casimir
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p. 2965 - 2972
(2007/10/03)
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- New highly selective inhibitors of class II fructose-1,6-bisphosphate aldolases
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Phosphoglycolo amidoxime and phosphoglycolo hydrazide, two new derivatives of phosphoglycolic acid, were synthesised and successfully tested as selective competitive inhibitors of class II FBP-aldolases.
- Fonvielle, Matthieu,Weber, Philippe,Dabkowska, Kasia,Therisod, Michel
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p. 2923 - 2926
(2007/10/03)
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- Structure-based mutagenesis approaches toward expanding the substrate specificity of D-2-deoxyribose-5-phosphate aldolase
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2-Deoxyribose-5-phosphate aldolase (DERA, EC 4.1.2.4) catalyzes the reversible aldol reaction between acetaldehyde and D-glyceraldehyde-3-phosphate to generate D-2-deoxyribose-5-phosphate. It is unique among the aldolases as it catalyzes the reversible asymmetric aldol addition reaction of two aldehydes. In order to expand the substrate scope and stereoselectivity of DERA, structure-based substrate design as well as site-specific mutation has been investigated. Using the 1.05 A crystal structure of DERA in complex with its natural substrate as a guide, five site-directed mutants were designed in order to improve its activity with the unnatural nonphosphorylated substrate, D-2-deoxyribose. Of these, the S238D variant exhibited a 2.5-fold improvement over the wild-type enzyme in the retroaldol reaction of 2-deoxyribose. Interestingly, this S238D mutant enzyme was shown to accept 3-azidopropinaldehyde as a substrate in a sequential asymmetric aldol reaction to form a deoxy-azidoethyl pyranose, which is a precursor to the corresponding lactone and the cholesterol-lowering agent Lipitor. This azidoaldehyde is not a substrate for the wild-type enzyme. Another structure-based design of new nonphosphorylated substrates was focused on the aldol reaction with inversion in enantioselectivity using the wild type or the S238D variant as the catalyst and 2-methyl-substituted aldehydes as substrates. An example was demonstrated in the asymmetric synthesis of a deoxypyranose as a new effective synthon for the total synthesis of epothilones. In addition, to facilitate the discovery of new enzymatic reactions, the engineered E. coli strain SELECT (Δace, adhC, DE3) was developed to be used in the future for selection of DERA variants with novel nonphosphorylated acceptor specificity.
- DeSantis, Grace,Liu, Junjie,Clark, David P.,Heine, Andreas,Wilson, Ian A.,Wong, Chi-Huey
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- Reactivities of some energy rich organic orthophosphates towards transition metal ion oxidants
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The orthophosphates, α- and β-glycerophosphates, react with HCrO-4 in HClO4 medium to give 1:1 intermediates which subsequently decomposes to give products of the reactions.The rate is directly proportional to +> in each reaction.Thermodynamic parameters associated with the equilibrium step and the activation parameters associated with the slowest step has been evaluated for each substrate.An attempt has been made to correlate the results obtained earlier with other transition metal ion oxidants.
- Gupta, Kalyan Kali Sen,Mahapatra, Ambikesh,Sanyal, Ankan
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p. 332 - 334
(2007/10/02)
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- Synthesis of Sugars by Aldolase-Catalyzed Condensation Reactions
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Dihydroxyacetone phosphate was prepared in 200-mmol scale from dihydroxyacetone by two procedures: reaction with phosphorus oxytrichloride and glycerol kinase catalyzed phosphorylation using ATP with in situ regeneration of ATP by phosphoenolpyruvate or acetyl phosphate.Dihydroxyacetone phosphate was converted to fructose 6-phosphate in 80percent yield by exposure to a mixture of co-immobilized triosephosphate isomerase and aldolase followed by acid hydrolysis of the condensation product fructose 1,6-bisphosphate.Fructose 6-phosphate was subsequently converted by chemical and enzymatic schemes into fructose, glucose 6-phosphate, and glucose.Practical procedures are described for the preparation of D- and L-glyceraldehyde 3-phosphate and for several hexoses labeled with 13C in the C-2 and C-2,5 positions.
- Wong, Chi-Huey,Whitesides, George M.
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p. 3199 - 3205
(2007/10/02)
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