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Anne K. Samland - One of the best experts on this subject based on the ideXlab platform.
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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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MJ0400 from Methanocaldococcus jannaschii exhibits fructose-1,6-bisphosphate aldolase activity
FEMS microbiology letters, 2008Co-Authors: Anne K. Samland, Mei Wang, Georg A SprengerAbstract:The central carbon metabolism is well investigated in bacteria, but this is not the case for archaea. MJ0400-His6 from Methanocaldococcus jannaschii catalyzes the cleavage of fructose-1,6-bisphosphate (FBP) to glyceraldehyde-3-phosphate and dihydroxyacetone phosphate with a V max of 33 mU mg−1 and a K m of 430 μM at 50 °C. MJ0400-His6 is inhibited competitively by Erythrose-4-Phosphate with a K i of 380 μM and displays heat stability with a half-life of c . 1 h at 100 °C. Hence, MJ0400 is the second gene encoding for an FBP aldolase in M. jannaschii . Previously, MJ0400 was shown to act as an 2-amino-3,7-dideoxy-d- threo -hept-6-ulosonic acid synthase. This indicates that MJ0400 is involved in both the carbon metabolism and the shikimate pathway in M. jannaschii .
Georg A Sprenger - One of the best experts on this subject based on the ideXlab platform.
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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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MJ0400 from Methanocaldococcus jannaschii exhibits fructose-1,6-bisphosphate aldolase activity
FEMS microbiology letters, 2008Co-Authors: Anne K. Samland, Mei Wang, Georg A SprengerAbstract:The central carbon metabolism is well investigated in bacteria, but this is not the case for archaea. MJ0400-His6 from Methanocaldococcus jannaschii catalyzes the cleavage of fructose-1,6-bisphosphate (FBP) to glyceraldehyde-3-phosphate and dihydroxyacetone phosphate with a V max of 33 mU mg−1 and a K m of 430 μM at 50 °C. MJ0400-His6 is inhibited competitively by Erythrose-4-Phosphate with a K i of 380 μM and displays heat stability with a half-life of c . 1 h at 100 °C. Hence, MJ0400 is the second gene encoding for an FBP aldolase in M. jannaschii . Previously, MJ0400 was shown to act as an 2-amino-3,7-dideoxy-d- threo -hept-6-ulosonic acid synthase. This indicates that MJ0400 is involved in both the carbon metabolism and the shikimate pathway in M. jannaschii .
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Aromatic Amino Acids
Amino Acid Biosynthesis ~ Pathways Regulation and Metabolic Engineering, 1Co-Authors: Georg A SprengerAbstract:Biosynthesis of the three aromatic amino acids (l-phenylalanine, l-trypto phan, l-tyrosine) and its regulation in Escherichia coli and corynebacteria are reviewed. The common aromatic biosynthetic pathway (shikimate pathway) starts with the condensation of phosphoenolpyruvate and Erythrose 4-Phosphate. Through six biosynthetic steps the pathway proceeds via shikimate to chorismate, from which the terminal pathways to tryptophan, phenylalanine and tyrosine branch. The first step in the common pathway is performed in E. coli by three isoenzymes, which are specifically feedback-inhibited by the three terminal products.
Sarah Schneider - One of the best experts on this subject based on the ideXlab platform.
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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.
Gunter Schneider - One of the best experts on this subject based on the ideXlab platform.
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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.
Tatjana Sandalova - One of the best experts on this subject based on the ideXlab platform.
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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.