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Accounts of Chemical Research May 2021Linear free energy relationships (LFERs) for substituent effects on reactions that proceed through similar transition states provide insight into transition state... (Review)
Review
Linear free energy relationships (LFERs) for substituent effects on reactions that proceed through similar transition states provide insight into transition state structures. A classical approach to the analysis of LFERs showed that differences in the slopes of Brønsted correlations for addition of substituted alkyl alcohols to ring-substituted 1-phenylethyl carbocations and to the β-galactopyranosyl carbocation intermediate of reactions catalyzed by β-galactosidase provide evidence that the enzyme catalyst modifies the curvature of the energy surface at the saddle point for the transition state for nucleophile addition. We have worked to generalize the use of LFERs in the determination of enzyme mechanisms. The defining property of enzyme catalysts is their specificity for binding the transition state with a much higher affinity than the substrate. Triosephosphate isomerase (TIM), orotidine 5'-monophosphate decarboxylase (OMPDC), and glycerol 3-phosphate dehydrogenase (GPDH) show effective catalysis of reactions of phosphorylated substrates and strong phosphite dianion activation of reactions of phosphodianion truncated substrates, with rate constants / (M s) and / (M s), respectively. Good linear logarithmic correlations, with a slope of 1.1, between these kinetic parameters determined for reactions catalyzed by five or more variant forms of each catalyst are observed, where the protein substitutions are mainly at side chains which function to stabilize the cage complex between the enzyme and substrate. This shows that the enzyme-catalyzed reactions of a whole substrate and substrate pieces proceed through transition states of similar structures. It provides support for the proposal that the dianion binding energy of whole phosphodianion substrates and of phosphite dianion is used to drive the conversion of these protein catalysts from flexible and entropically rich ground states to stiff and catalytically active Michaelis complexes that show the same activity toward catalysis of the reactions of whole and phosphodianion truncated substrates. There is a good linear correlation, with a slope of 0.73, between values of the dissociation constants log for release of the transition state analog phosphoglycolate (PGA) trianion and log / for isomerization of GAP for wild-type and variants of TIM. This correlation shows that the substituted amino acid side chains act to stabilize the complex between TIM and the PGA trianion and that . 70% of this stabilization is observed at the transition state for substrate deprotonation. The correlation provides evidence that these side chains function to enhance the basicity of the E165 side chain of TIM, which deprotonates the bound carbon acid substrate. There is a good linear correlation, with a slope of 0.74, between the values of Δ and Δ° determined by electron valence bond (EVB) calculations to model deprotonation of dihydroxyacetone phosphate (DHAP) in water and when bound to wild-type and variant forms of TIM to form the enediolate reaction intermediate. This correlation provides evidence that the stabilizing interactions of the transition state for TIM-catalyzed deprotonation of DHAP are optimized by placement of amino acid side chains in positions that provide for the maximum stabilization of the charged reaction intermediate, relative to the neutral substrate.
Topics: Humans; Models, Molecular; Thermodynamics; Triose-Phosphate Isomerase
PubMed: 33939414
DOI: 10.1021/acs.accounts.1c00147 -
Biochimica Et Biophysica Acta.... Jun 2021Phosphatidic acid biosynthesis represents the initial part of de novo formation of all glycerophospholipids (membrane lipids) as well as triacylglycerols (storage... (Review)
Review
Phosphatidic acid biosynthesis represents the initial part of de novo formation of all glycerophospholipids (membrane lipids) as well as triacylglycerols (storage lipids), and is thus the centerpiece of glycerolipid metabolism. The universal route of phosphatidic acid biosynthesis starts from the precursor glycerol-3-phosphate and comprises two consecutive acylation reactions which are catalyzed by a glycerol-3-phosphate acyltransferase and a 1-acyl glycerol-3-phosphate acyltransferase. In addition, yeast and mammals harbor a set of enzymes which can synthesize phosphatidic acid from the precursor dihydroxyacetone phosphate. In the present review our current knowledge about enzymes contributing to phosphatidic acid biosynthesis in the invaluable model organism yeast, Saccharomyces cerevisiae, is summarized. A special focus is laid upon the regulation and the localization of these enzymes. Furthermore, research needs for a deeper insight into the high complexity of phosphatidic acid biosynthesis and consequently the entire lipid metabolic network is presented.
Topics: Phosphatidic Acids; Saccharomyces cerevisiae
PubMed: 33610760
DOI: 10.1016/j.bbalip.2021.158907 -
Molecules (Basel, Switzerland) Mar 20231,3-dihydroxyacetone (DHA) is an underrated bio-based synthon, with a broad range of reactivities. It is produced for the revalorization of glycerol, a major... (Review)
Review
1,3-dihydroxyacetone (DHA) is an underrated bio-based synthon, with a broad range of reactivities. It is produced for the revalorization of glycerol, a major side-product of the growing biodiesel industry. The overwhelming majority of DHA produced worldwide is intended for application as a self-tanning agent in cosmetic formulations. This review provides an overview of the discovery, physical and chemical properties of DHA, and of its industrial production routes from glycerol. Microbial fermentation is the only industrial-scaled route but advances in electrooxidation and aerobic oxidation are also reported. This review focuses on the plurality of reactivities of DHA to help chemists interested in bio-based building blocks see the potential of DHA for this application. The handling of DHA is delicate as it can undergo dimerization as well as isomerization reactions in aqueous solutions at room temperature. DHA can also be involved in further side-reactions, yielding original side-products, as well as compounds of interest. If this peculiar reactivity was harnessed, DHA could help address current sustainability challenges encountered in the synthesis of speciality polymers, ranging from biocompatible polymers to innovative polymers with cutting-edge properties and improved biodegradability.
Topics: Dihydroxyacetone; Glycerol; Fermentation; Oxidation-Reduction; Cosmetics
PubMed: 36985712
DOI: 10.3390/molecules28062724 -
Frontiers in Genetics 2022Trehalose is commonly used as an impermeable cryoprotectant for cryopreservation of cells, but its cryoprotective mechanism has now not but been determined. This study...
Trehalose is commonly used as an impermeable cryoprotectant for cryopreservation of cells, but its cryoprotective mechanism has now not but been determined. This study investigated the cryopreservation impact of trehalose on buck semen cryopreservation and finished metabolic profiling of freeze-thawed media by way of the GC-MS-based metabolomics for the first time. Metabolic pattern recognition and metabolite identification by means of principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA) and metabolic pathway topology analysis revealed the results of trehalose on buck sperm metabolism at some point of cryopreservation. The results confirmed that trehalose drastically progressed sperm motility parameters and structural integrity after thawing. PCA and PLS-DA analysis discovered that the metabolic patterns of the freezing-thawing media of buck semen cryopreserved with trehalose (T group) or without trehalose (G group, Control) were certainly separated. Using screening conditions of VIP >1.5 and vaule <0.05, a total of 48 differential metabolites have been recognized, whithin l-isoleucine, L-leucine, L-threonine, and dihydroxyacetone were notably enriched in valine, leucine and isoleucine biosynthesis, glycerolipid metabolism, and aminoacyl-tRNA biosynthesis pathways. In brief, trehalose can efficiently improve membrane structural integrity and motion parameters in buck sperm after thawing, and it exerts a cryoprotective impact with the aid of changing sperm amino acid synthesis and the glycerol metabolism pathway.
PubMed: 35991557
DOI: 10.3389/fgene.2022.938622 -
Microbial Cell Factories Jun 2021Klebsiella pneumoniae is a bacterium that can be used as producer for numerous chemicals. Glycerol can be catabolised by K. pneumoniae and dihydroxyacetone is an...
BACKGROUND
Klebsiella pneumoniae is a bacterium that can be used as producer for numerous chemicals. Glycerol can be catabolised by K. pneumoniae and dihydroxyacetone is an intermediate of this catabolism pathway. Here dihydroxyacetone and glycerol were produced from glucose by this bacterium based a redirected glycerol catabolism pathway.
RESULTS
tpiA, encoding triosephosphate isomerase, was knocked out to block the further catabolism of dihydroxyacetone phosphate in the glycolysis. After overexpression of a Corynebacterium glutamicum dihydroxyacetone phosphate dephosphorylase (hdpA), the engineered strain produced remarkable levels of dihydroxyacetone (7.0 g/L) and glycerol (2.5 g/L) from glucose. Further increase in product formation were obtained by knocking out gapA encoding an iosenzyme of glyceraldehyde 3-phosphate dehydrogenase. There are two dihydroxyacetone kinases in K. pneumoniae. They were both disrupted to prevent an inefficient reaction cycle between dihydroxyacetone phosphate and dihydroxyacetone, and the resulting strains had a distinct improvement in dihydroxyacetone and glycerol production. pH 6.0 and low air supplement were identified as the optimal conditions for dihydroxyacetone and glycerol production by K, pneumoniae ΔtpiA-ΔDHAK-hdpA. In fed batch fermentation 23.9 g/L of dihydroxyacetone and 10.8 g/L of glycerol were produced after 91 h of cultivation, with the total conversion ratio of 0.97 mol/mol glucose.
CONCLUSIONS
This study provides a novel and highly efficient way of dihydroxyacetone and glycerol production from glucose.
Topics: Dihydroxyacetone; Dihydroxyacetone Phosphate; Diphosphoglyceric Acids; Fermentation; Genes, Bacterial; Glucose; Glyceraldehyde 3-Phosphate; Glyceraldehyde-3-Phosphate Dehydrogenases; Glycerol; Hydrogen-Ion Concentration; Klebsiella pneumoniae; Metabolic Engineering; Metabolic Networks and Pathways; Phosphotransferases (Alcohol Group Acceptor); Thermodynamics
PubMed: 34187467
DOI: 10.1186/s12934-021-01608-0 -
Journal of Oral Microbiology 2022The etiology of dental caries remains poorly understood. With the advent of next-generation sequencing, a number of studies have focused on the microbial ecology of the...
BACKGROUND
The etiology of dental caries remains poorly understood. With the advent of next-generation sequencing, a number of studies have focused on the microbial ecology of the disease. However, taxonomic associations with caries have not been consistent. Researchers have also pursued function-centric studies of the caries microbial communities aiming to identify consistently conserved functional pathways. A major question is whether changes in microbiome are a cause or a consequence of the disease. Thus, there is a critical need to define conserved functional signatures at the onset of dental caries.
METHODS
Since it is unethical to induce carious lesions clinically, we developed an innovative longitudinal model integrated with the advanced non-invasive multiphoton second harmonic generation bioimaging to spot the very early signs of dental caries, combined with 16S rRNA short amplicon sequencing and liquid chromatography-mass spectrometry-based targeted metabolomics.
FINDINGS
For the first time, we induced longitudinally monitored caries lesions validated with the scanning electron microscope. Consequently, we spotted the caries onset and, associated with it, distinguished five differentiating metabolites - Lactate, Pyruvate, Dihydroxyacetone phosphate, Glyceraldehyde 3-phosphate (upregulated) and Fumarate (downregulated). Those metabolites co-occurred with certain bacterial taxa; , and , regardless of the abundance of other taxa.
INTERPRETATION
These findings are crucial for understanding the etiology and dynamics of dental caries, and devising targeted interventions to prevent disease progression.
PubMed: 36189437
DOI: 10.1080/20002297.2022.2123624 -
Biochemistry Jun 2023Four catalytic amino acids at triosephosphate isomerase (TIM) are highly conserved: N11, K13, H95, and E167. Asparagine 11 is the last of these to be characterized in...
Four catalytic amino acids at triosephosphate isomerase (TIM) are highly conserved: N11, K13, H95, and E167. Asparagine 11 is the last of these to be characterized in mutagenesis studies. The ND2 side chain atom of N11 is hydrogen bonded to the O-1 hydroxyl of enzyme-bound dihydroxyacetone phosphate (DHAP), and it sits in an extended chain of hydrogen-bonded side chains that includes T75' from the second subunit. The N11A variants of wild-type TIM from (TIM) and (TIM) undergo dissociation from the dimer to monomer under our assay conditions. Values of = 8 × 10 and 1 × 10 M, respectively, were determined for the conversion of monomeric N11A TIM and TIM into their homodimers. The N11A substitution at the variant of TIM previously stabilized by the E65Q substitution gives the N11A/E65Q variant that is stable to dissociation under our assay conditions. The X-ray crystal structure of N11A/E65Q TIM shows an active site that is essentially superimposable on that for wild-type TIM, which also has a glutamine at position 65. A comparison of the kinetic parameters for E65Q TIM and N11A/E65Q TIM-catalyzed reactions of ()-glyceraldehyde 3-phosphate (GAP) and (DHAP) shows that the N11A substitution results in a (13-14)-fold decrease in / for substrate isomerization and a similar decrease in for DHAP but only a 2-fold decrease in for GAP.
Topics: Triose-Phosphate Isomerase; Catalysis; Amino Acids; Hydrogen
PubMed: 37162263
DOI: 10.1021/acs.biochem.3c00133 -
The Journal of Biological Chemistry Nov 2023Sulfoquinovose (SQ, 6-deoxy-6-sulfoglucose) is a sulfosugar that is the anionic head group of plant, algal, and cyanobacterial sulfolipids: sulfoquinovosyl...
Sulfoquinovose (SQ, 6-deoxy-6-sulfoglucose) is a sulfosugar that is the anionic head group of plant, algal, and cyanobacterial sulfolipids: sulfoquinovosyl diacylglycerols. SQ is produced within photosynthetic tissues, forms a major terrestrial reservoir of biosulfur, and is an important species within the biogeochemical sulfur cycle. A major pathway for SQ breakdown is the sulfoglycolytic Embden-Meyerhof-Parnas pathway, which involves cleavage of the 6-carbon chain of the intermediate sulfofructose-1-phosphate (SFP) into dihydroxyacetone and sulfolactaldehyde, catalyzed by class I or II SFP aldolases. While the molecular basis of catalysis is understood for class I SFP aldolases, comparatively little is known about class II SFP aldolases. Here, we report the molecular architecture and biochemical basis of catalysis of two metal-dependent class II SFP aldolases from Hafnia paralvei and Yersinia aldovae. 3D X-ray structures of complexes with substrate SFP and product dihydroxyacetone phosphate reveal a dimer-of-dimers (tetrameric) assembly, the sulfonate-binding pocket, two metal-binding sites, and flexible loops that are implicated in catalysis. Both enzymes were metal-dependent and exhibited high K values for SFP, consistent with their role in a unidirectional nutrient acquisition pathway. Bioinformatic analysis identified a range of sulfoglycolytic Embden-Meyerhof-Parnas gene clusters containing class I/II SFP aldolases. The class I and II SFP aldolases have mututally exclusive occurrence within Actinobacteria and Firmicutes phyla, respectively, while both classes of enzyme occur within Proteobacteria. This work emphasizes the importance of SQ as a nutrient for diverse bacterial phyla and the different chemical strategies they use to harvest carbon from this sulfosugar.
Topics: Aldehyde-Lyases; Carbon; Fructose-Bisphosphate Aldolase; Metals; Phosphates
PubMed: 37838169
DOI: 10.1016/j.jbc.2023.105338 -
Cells Mar 2022Upregulation of glycolysis, induction of epithelial-mesenchymal transition (EMT) and macroautophagy (hereafter autophagy), are phenotypic changes that occur in tumor... (Review)
Review
Upregulation of glycolysis, induction of epithelial-mesenchymal transition (EMT) and macroautophagy (hereafter autophagy), are phenotypic changes that occur in tumor cells, in response to similar stimuli, either tumor cell-autonomous or from the tumor microenvironment. Available evidence, herein reviewed, suggests that glycolysis can play a causative role in the induction of EMT and autophagy in tumor cells. Thus, glycolysis has been shown to induce EMT and either induce or inhibit autophagy. Glycolysis-induced autophagy occurs both in the presence (glucose starvation) or absence (glucose sufficiency) of metabolic stress. In order to explain these, in part, contradictory experimental observations, we propose that in the presence of stimuli, tumor cells respond by upregulating glycolysis, which will then induce EMT and inhibit autophagy. In the presence of stimuli and glucose starvation, upregulated glycolysis leads to adenosine monophosphate-activated protein kinase (AMPK) activation and autophagy induction. In the presence of stimuli and glucose sufficiency, upregulated glycolytic enzymes (e.g., aldolase or glyceraldehyde 3-phosphate dehydrogenase) or decreased levels of glycolytic metabolites (e.g., dihydroxyacetone phosphate) may mimic a situation of metabolic stress (herein referred to as "pseudostarvation"), leading, directly or indirectly, to AMPK activation and autophagy induction. We also discuss possible mechanisms, whereby glycolysis can induce a mixed mesenchymal/autophagic phenotype in tumor cells. Subsequently, we address unresolved problems in this field and possible therapeutic consequences.
Topics: AMP-Activated Protein Kinases; Autophagy; Epithelial-Mesenchymal Transition; Glucose; Glycolysis
PubMed: 35326492
DOI: 10.3390/cells11061041 -
Foods (Basel, Switzerland) Jul 2021Soil properties in the foraging range of honeybees influence honey composition. We aimed to determine relationships between the antimicrobial properties of New Zealand...
Soil properties in the foraging range of honeybees influence honey composition. We aimed to determine relationships between the antimicrobial properties of New Zealand mānuka () honey and elemental concentrations in the honey, plants, and soils. We analyzed soils, plants, and fresh mānuka honey samples from the Wairarapa region of New Zealand for the chemical elements and the antimicrobial activity of the honey as indicated by methylglyoxal (MGO) and dihydroxyacetone (DHA). There were significant negative correlations between honey MGO and the concentrations of Mn, Cu, Mg, S, Na, Ba, K, Zn, and Al. These elements may provide a low-cost means of assessing mānuka honey quality. For individual elements, except for K, there were no correlations between the honeys, plants, and soils. Soil nitrate concentrations were negatively correlated with concentrations of MGO and DHA in the honey, which implies that soil fertility may be a determiner of mānuka honey quality.
PubMed: 34359540
DOI: 10.3390/foods10071670