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Carlos Maria Figueroa - One of the best experts on this subject based on the ideXlab platform.
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The Crystal Structure of Nitrosomonas europaea Sucrose Synthase Reveals Critical Conformational Changes and Insights into Sucrose Metabolism in Prokaryotes
Journal of Bacteriology, 2015Co-Authors: Matias Damian Asencion Diez, Miguel A Ballicora, Alberto A Iglesias, Carlos Maria Figueroa, Matías Machtey, Dali LiuAbstract:In this paper we report the first crystal structure of a prokaryotic Sucrose Synthase from the nonphotosynthetic bacterium Nitrosomonas europaea. The obtained structure was in an open form, whereas the only other available structure, from the plant Arabidopsis thaliana, was in a closed conformation. Comparative structural analysis revealed a “hinge-latch” combination, which is critical to transition between the open and closed forms of the enzyme. The N. europaea Sucrose Synthase shares the same fold as the GT-B family of the retaining glycosyltransferases. In addition, a triad of conserved homologous catalytic residues in the family was shown to be functionally critical in the N. europaea Sucrose Synthase (Arg567, Lys572, and Glu663). This implies that Sucrose Synthase shares not only a common origin with the GT-B family but also a similar catalytic mechanism. The enzyme preferred transferring glucose from ADP-glucose rather than UDP-glucose like the eukaryotic counterparts. This predicts that these prokaryotic organisms have a different Sucrose metabolic scenario from plants. Nucleotide preference determines where the glucose moiety is targeted after Sucrose is degraded. IMPORTANCE We obtained biochemical and structural evidence of Sucrose metabolism in nonphotosynthetic bacteria. Until now, only Sucrose Synthases from photosynthetic organisms have been characterized. Here, we provide the crystal structure of the Sucrose Synthase from the chemolithoautotroph N. europaea. The structure supported that the enzyme functions with an open/close induced fit mechanism. The enzyme prefers as the substrate adenine-based nucleotides rather than uridine-based like the eukaryotic counterparts, implying a strong connection between Sucrose and glycogen metabolism in these bacteria. Mutagenesis data showed that the catalytic mechanism must be conserved not only in Sucrose Synthases but also in all other retaining GT-B glycosyltransferases.
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the unique nucleotide specificity of the Sucrose Synthase from thermosynechococcus elongatus
FEBS Letters, 2013Co-Authors: Matias Damian Asencion Diez, Sheila Mcewen, Graciela Lidia Salerno, Alberto A Iglesias, Carlos Maria Figueroa, Misty L Kuhn, Miguel A BallicoraAbstract:Sucrose Synthase catalyzes the reversible conversion of Sucrose and UDP into fructose and UDP-glucose. In filamentous cyanobacteria, the Sucrose cleavage direction plays a key physiological function in carbon metabolism, nitrogen fixation, and stress tolerance. In unicellular strains, the function of Sucrose Synthase has not been elucidated. We report a detailed biochemical characterization of Sucrose Synthase from Thermosynechococcus elongatus after the gene was artificially synthesized for optimal expression in Escherichia coli. The homogeneous recombinant Sucrose Synthase was highly specific for ADP as substrate, constituting the first one with this unique characteristic, and strongly suggesting an interaction between Sucrose and glycogen metabolism.
Fernando Luiz Finger - One of the best experts on this subject based on the ideXlab platform.
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Colocalization of Sucrose Synthase expression and Sucrose storage in the sugarbeet taproot indicates a potential role for Sucrose catabolism in Sucrose accumulation.
Journal of Plant Physiology, 2019Co-Authors: Karen Klotz Fugate, John D. Eide, Daniel N. Martins, Edward L. Deckard, Michael A. Grusak, Fernando Luiz FingerAbstract:Abstract Sucrose metabolism is believed to have a central role in promoting sink strength and Sucrose storage in the sugarbeet taproot. How Sucrose accumulation is increased by Sucrose-degrading enzymes, however, is a paradox. To elucidate roles for Sucrose-degrading activities in Sucrose accumulation, relationships between the intercellular location of Sucrose-catabolizing enzymes and sites of Sucrose accumulation were determined in the sugarbeet taproot. Sucrose storage was evident in parenchyma cells of the outer cortex, rays, and rings of parenchyma tissue, but was absent in phloem, the vascular cambium, cells surrounding these tissues, or cells surrounding xylem. Sucrose Synthase, which was primarily responsible for Sucrose catabolism throughout the taproot, was expressed in similar cell and tissue types to those accumulating Sucrose. Colocalization of Sucrose Synthase with Sucrose accumulation, as well as Sucrose Synthase localization near the tonoplast, suggests a role for the enzyme in generating metabolic energy to fuel Sucrose sequestration in the vacuole. Localization near the plasma membrane also suggests a role for Sucrose Synthase in supplying substrates for cell wall biosynthesis. By utilizing Sucrose for ATP or cell wall biosynthesis, Sucrose Synthase likely maintains the source-to-sink Sucrose gradient that drives Sucrose transport into the root, thereby promoting sugarbeet root sink strength.
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Characterization of two Sucrose Synthase isoforms in sugarbeet root
Plant Physiology and Biochemistry, 2003Co-Authors: Karen L. Klotz, Fernando Luiz Finger, Weilin L. ShelverAbstract:Abstract Two Sucrose Synthase isoforms (EC 2.4.1.13) have been identified in developing sugarbeet (Beta vulgaris L.) roots. To aid in understanding the physiological significance of these multiple Sucrose Synthase isoforms, the two isoforms were partially purified and some of their physical and kinetic properties determined. Both isoforms were tetrameric proteins with native molecular masses of 320 kDa. The isoforms exhibited similar kinetic properties as well as similar changes in activity in response to changes in temperature. The isoforms differed, however, in their subunit composition. Sucrose Synthase isoform I (SuSyI) was composed of two 84 kDa subunits and two 86 kDa subunits. Sucrose Synthase isoform II (SuSyII) was a homotetramer with a subunit size of 86 kDa. The amino acid composition of the two subunits was similar, although differences in alanine, glycine, isoleucine and lysine content were noted. The activity of the two isoforms differed in response to varying pH conditions. The optimum pH for Sucrose cleaving activity was observed at pH 6.0 and 6.5 for SuSyI and SuSyII, respectively. The optimum pH for Sucrose synthesizing activity occurred at pH 7.5 and 7.0 for SuSyI and SuSyII, respectively. The observed differences in subunit composition and reactivity at different pH values suggest that multiple isoforms of Sucrose Synthase may provide a mechanism to regulate Sucrose metabolism in sugarbeet root by differential regulation of expression of the two isoforms and modulation of their activity by changes in cellular pH.
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Contribution of Invertase and Sucrose Synthase Isoforms to Sucrose Catabolism in Developing Sugarbeet Roots
Journal of Sugarbeet Research, 2002Co-Authors: Karen L. Klotzl, Fernando Luiz FingerAbstract:Isoforms of the major sucrolytic enzymes in sugarbeet were identified and their developmental expressions of activity were determined with respect to root growth and carbohydrate accumulation. Sugarbeet roots contained at least seven different sucrolytic activities throughout their development. Two soluble acid invertase isoforms, an insoluble acid invertase activity, two alkaline invertase isoforms and two Sucrose Synthase isoforms were identified. Each enzyme isoform exhibited a unique pattern of developmental expression. Soluble and insoluble acid invertase activities were the predominant sucrolytic activities in young roots and declined rapidly as roots aged. Soluble acid invertase activity was due primarily to the activity of a single isoform, although a second minor isoform was evident in roots of seedlings. High soluble and insoluble acid invertase activities occurred concurrently with a rapid relative growth rate, high glucose concentration and minimal Sucrose accumulation. Sucrose Synthase was the major sucrolytic activity during most of root development and was the predominant sucrolytic activity during the period in which nearly all Sucrose accumulation and enlargement of the taproot occurred. Sucrose Synthase activity correlated highly with absolute growth rate of the root. One Sucrose Synthase isoform was present throughout development. A second isoform became evident as roots approached maturity. Alkaline invertase activity was present at low, relatively constant activities at all but the earliest stages of development and was due to two isoforms whose contribution to total alkaline invertase activity changed as roots matured. The presence of multiple, differentially regulated, sucrolytic enzymes seemingly allows control of Sucrose catabolism to balance the metabolic needs of the growing sugarbeet root with its function as a Sucrose storage organ.
Miguel A Ballicora - One of the best experts on this subject based on the ideXlab platform.
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The Crystal Structure of Nitrosomonas europaea Sucrose Synthase Reveals Critical Conformational Changes and Insights into Sucrose Metabolism in Prokaryotes
Journal of Bacteriology, 2015Co-Authors: Matias Damian Asencion Diez, Miguel A Ballicora, Alberto A Iglesias, Carlos Maria Figueroa, Matías Machtey, Dali LiuAbstract:In this paper we report the first crystal structure of a prokaryotic Sucrose Synthase from the nonphotosynthetic bacterium Nitrosomonas europaea. The obtained structure was in an open form, whereas the only other available structure, from the plant Arabidopsis thaliana, was in a closed conformation. Comparative structural analysis revealed a “hinge-latch” combination, which is critical to transition between the open and closed forms of the enzyme. The N. europaea Sucrose Synthase shares the same fold as the GT-B family of the retaining glycosyltransferases. In addition, a triad of conserved homologous catalytic residues in the family was shown to be functionally critical in the N. europaea Sucrose Synthase (Arg567, Lys572, and Glu663). This implies that Sucrose Synthase shares not only a common origin with the GT-B family but also a similar catalytic mechanism. The enzyme preferred transferring glucose from ADP-glucose rather than UDP-glucose like the eukaryotic counterparts. This predicts that these prokaryotic organisms have a different Sucrose metabolic scenario from plants. Nucleotide preference determines where the glucose moiety is targeted after Sucrose is degraded. IMPORTANCE We obtained biochemical and structural evidence of Sucrose metabolism in nonphotosynthetic bacteria. Until now, only Sucrose Synthases from photosynthetic organisms have been characterized. Here, we provide the crystal structure of the Sucrose Synthase from the chemolithoautotroph N. europaea. The structure supported that the enzyme functions with an open/close induced fit mechanism. The enzyme prefers as the substrate adenine-based nucleotides rather than uridine-based like the eukaryotic counterparts, implying a strong connection between Sucrose and glycogen metabolism in these bacteria. Mutagenesis data showed that the catalytic mechanism must be conserved not only in Sucrose Synthases but also in all other retaining GT-B glycosyltransferases.
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the unique nucleotide specificity of the Sucrose Synthase from thermosynechococcus elongatus
FEBS Letters, 2013Co-Authors: Matias Damian Asencion Diez, Sheila Mcewen, Graciela Lidia Salerno, Alberto A Iglesias, Carlos Maria Figueroa, Misty L Kuhn, Miguel A BallicoraAbstract:Sucrose Synthase catalyzes the reversible conversion of Sucrose and UDP into fructose and UDP-glucose. In filamentous cyanobacteria, the Sucrose cleavage direction plays a key physiological function in carbon metabolism, nitrogen fixation, and stress tolerance. In unicellular strains, the function of Sucrose Synthase has not been elucidated. We report a detailed biochemical characterization of Sucrose Synthase from Thermosynechococcus elongatus after the gene was artificially synthesized for optimal expression in Escherichia coli. The homogeneous recombinant Sucrose Synthase was highly specific for ADP as substrate, constituting the first one with this unique characteristic, and strongly suggesting an interaction between Sucrose and glycogen metabolism.
Matias Damian Asencion Diez - One of the best experts on this subject based on the ideXlab platform.
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The Crystal Structure of Nitrosomonas europaea Sucrose Synthase Reveals Critical Conformational Changes and Insights into Sucrose Metabolism in Prokaryotes
Journal of Bacteriology, 2015Co-Authors: Matias Damian Asencion Diez, Miguel A Ballicora, Alberto A Iglesias, Carlos Maria Figueroa, Matías Machtey, Dali LiuAbstract:In this paper we report the first crystal structure of a prokaryotic Sucrose Synthase from the nonphotosynthetic bacterium Nitrosomonas europaea. The obtained structure was in an open form, whereas the only other available structure, from the plant Arabidopsis thaliana, was in a closed conformation. Comparative structural analysis revealed a “hinge-latch” combination, which is critical to transition between the open and closed forms of the enzyme. The N. europaea Sucrose Synthase shares the same fold as the GT-B family of the retaining glycosyltransferases. In addition, a triad of conserved homologous catalytic residues in the family was shown to be functionally critical in the N. europaea Sucrose Synthase (Arg567, Lys572, and Glu663). This implies that Sucrose Synthase shares not only a common origin with the GT-B family but also a similar catalytic mechanism. The enzyme preferred transferring glucose from ADP-glucose rather than UDP-glucose like the eukaryotic counterparts. This predicts that these prokaryotic organisms have a different Sucrose metabolic scenario from plants. Nucleotide preference determines where the glucose moiety is targeted after Sucrose is degraded. IMPORTANCE We obtained biochemical and structural evidence of Sucrose metabolism in nonphotosynthetic bacteria. Until now, only Sucrose Synthases from photosynthetic organisms have been characterized. Here, we provide the crystal structure of the Sucrose Synthase from the chemolithoautotroph N. europaea. The structure supported that the enzyme functions with an open/close induced fit mechanism. The enzyme prefers as the substrate adenine-based nucleotides rather than uridine-based like the eukaryotic counterparts, implying a strong connection between Sucrose and glycogen metabolism in these bacteria. Mutagenesis data showed that the catalytic mechanism must be conserved not only in Sucrose Synthases but also in all other retaining GT-B glycosyltransferases.
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the unique nucleotide specificity of the Sucrose Synthase from thermosynechococcus elongatus
FEBS Letters, 2013Co-Authors: Matias Damian Asencion Diez, Sheila Mcewen, Graciela Lidia Salerno, Alberto A Iglesias, Carlos Maria Figueroa, Misty L Kuhn, Miguel A BallicoraAbstract:Sucrose Synthase catalyzes the reversible conversion of Sucrose and UDP into fructose and UDP-glucose. In filamentous cyanobacteria, the Sucrose cleavage direction plays a key physiological function in carbon metabolism, nitrogen fixation, and stress tolerance. In unicellular strains, the function of Sucrose Synthase has not been elucidated. We report a detailed biochemical characterization of Sucrose Synthase from Thermosynechococcus elongatus after the gene was artificially synthesized for optimal expression in Escherichia coli. The homogeneous recombinant Sucrose Synthase was highly specific for ADP as substrate, constituting the first one with this unique characteristic, and strongly suggesting an interaction between Sucrose and glycogen metabolism.
Karen L. Klotz - One of the best experts on this subject based on the ideXlab platform.
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Wounding, anoxia and cold induce sugarbeet Sucrose Synthase transcriptional changes that are unrelated to protein expression and activity.
Journal of Plant Physiology, 2008Co-Authors: Karen L. Klotz, Darrin M. HaagensonAbstract:Wounding, anoxia, and cold are often encountered during crop production and postharvest storage of plant products. Although the effect of these stresses on the expression of Sucrose Synthase, a key enzyme in the carbon metabolism of many storage organs, has been investigated in several starch-accumulating plant organs, little information on their effect on Sucrose Synthase expression in Sucrose-storing organs is available. To determine the effect of wounding, anoxia and cold on a Sucrose-storing organ, sugarbeet (Beta vulgaris) roots were wounded, subjected to anoxic conditions, or exposed to cold temperatures, and transcript and protein levels for the organ's two Sucrose Synthase genes (SBSS1 and SBSS2) and Sucrose Synthase enzyme activity were determined during 24 h and 7 d time course experiments. Wounding, anoxia and cold were associated with several-fold changes in Sucrose Synthase transcript levels. SBSS1 transcript levels were elevated in wounded, anoxic and cold-treated roots; SBSS2 transcript levels were elevated in response to wounding, cold, and short exposures (3–12 h) to anoxic conditions and reduced in roots exposed to anoxic conditions for more than 24 h. SBSS1 and SBSS2 protein levels, however, exhibited little change in stressed roots, even after 7 d. Enzyme activity was also relatively unchanged in stressed roots, except for small activity differences of 1–2 d duration that were unrelated to transcriptional changes. The disparity between transcript levels, protein abundance and enzyme activity indicate that SBSS1 and SBSS2 expression in response to wounding, anoxia and cold may be regulated by post-transcriptional mechanisms. The unresponsiveness of Sucrose Synthase protein levels or enzyme activity to wounding, anoxia and cold questions the importance of this enzyme to stress responses in sugarbeet root.
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Sugarbeet Sucrose Synthase genes differ in organ-specific and developmental expression
Journal of Plant Physiology, 2006Co-Authors: Darrin M. Haagenson, Karen L. Klotz, J. Mitchell McgrathAbstract:A full-length Sucrose Synthase (SBSS2) cDNA clone was isolated from sugarbeet. Comparison of its composition and organ-specific and developmental expression with a previously isolated sugarbeet Sucrose Synthase gene (SBSS1) revealed distinct differences between the two genes. The two genes share 80% similarity in deduced amino acid sequence but belong to different Sucrose Synthase subclasses based on phylogenic analysis. Both Sucrose Synthases were highly expressed in roots, and had low levels of expression in leaf tissue. Transcript abundance of SBSS2, relative to SBSS1, was greater in young vegetative and floral tissues, and reduced in mature vegetative tissues. The organ-specific and developmental expression of SBSS1 and SBSS2 proteins was similar to SBSS1 and SBSS2 transcript levels, although developmental changes in protein abundance lagged transcriptional changes by many weeks. The similarities and differences in transcript and protein abundance suggest that both transcriptional and post-transcriptional regulatory mechanisms are likely to contribute to Sucrose Synthase expression in sugarbeet.
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Characterization of two Sucrose Synthase isoforms in sugarbeet root
Plant Physiology and Biochemistry, 2003Co-Authors: Karen L. Klotz, Fernando Luiz Finger, Weilin L. ShelverAbstract:Abstract Two Sucrose Synthase isoforms (EC 2.4.1.13) have been identified in developing sugarbeet (Beta vulgaris L.) roots. To aid in understanding the physiological significance of these multiple Sucrose Synthase isoforms, the two isoforms were partially purified and some of their physical and kinetic properties determined. Both isoforms were tetrameric proteins with native molecular masses of 320 kDa. The isoforms exhibited similar kinetic properties as well as similar changes in activity in response to changes in temperature. The isoforms differed, however, in their subunit composition. Sucrose Synthase isoform I (SuSyI) was composed of two 84 kDa subunits and two 86 kDa subunits. Sucrose Synthase isoform II (SuSyII) was a homotetramer with a subunit size of 86 kDa. The amino acid composition of the two subunits was similar, although differences in alanine, glycine, isoleucine and lysine content were noted. The activity of the two isoforms differed in response to varying pH conditions. The optimum pH for Sucrose cleaving activity was observed at pH 6.0 and 6.5 for SuSyI and SuSyII, respectively. The optimum pH for Sucrose synthesizing activity occurred at pH 7.5 and 7.0 for SuSyI and SuSyII, respectively. The observed differences in subunit composition and reactivity at different pH values suggest that multiple isoforms of Sucrose Synthase may provide a mechanism to regulate Sucrose metabolism in sugarbeet root by differential regulation of expression of the two isoforms and modulation of their activity by changes in cellular pH.
