The Experts below are selected from a list of 180 Experts worldwide ranked by ideXlab platform
Mahdi M. Abu-omar - One of the best experts on this subject based on the ideXlab platform.
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Role of the second coordination sphere residue tyrosine 179 in substrate affinity and catalytic activity of Phenylalanine hydroxylase
JBIC Journal of Biological Inorganic Chemistry, 2004Co-Authors: Jérôme Zoidakis, Alon Volner, Kim Vu, Mahdi M. Abu-omarAbstract:Phenylalanine hydroxylase converts Phenylalanine to tyrosine utilizing molecular oxygen and tetrahydropterin as a cofactor, and belongs to the aromatic amino acid hydroxylases family. The catalytic domains of these enzymes are structurally similar. According to recent crystallographic studies, residue Tyr179 in Chromobacterium violaceum Phenylalanine hydroxylase is located in the active site and its hydroxyl oxygen is 5.1 Å from the iron, where it has been suggested to play a role in positioning the pterin cofactor. To determine the catalytic role of this residue, the point mutants Y179F and Y179A of Phenylalanine hydroxylase were prepared and characterized. Both mutants displayed comparable stability and metal binding to the native enzyme, as determined by their melting temperatures in the presence and absence of iron. The catalytic activity ( k _cat) of the Y179F and Y179A proteins was lower than wild-type Phenylalanine hydroxylase by an order of magnitude, suggesting that the hydroxyl group of Tyr179 plays a role in the rate-determining step in catalysis. The K _M values for different tetrahydropterin cofactors and Phenylalanine were decreased by a factor of 3–4 in the Y179F mutant. However, the K _M values for different pterin cofactors were slightly higher in the Y179A mutant than those measured for the wild-type enzyme, and, more significantly, the K _M value for Phenylalanine was increased by 10-fold in the Y179A mutant. By the criterion of k _cat/ K _Phe, the Y179F and Y179A mutants display 10% and 1%, respectively, of the activity of wild-type Phenylalanine hydroxylase. These results are consistent with Tyr179 having a pronounced role in binding Phenylalanine but a secondary effect in the formation of the hydroxylating species. In conjunction with recent crystallographic analyses of a ternary complex of Phenylalanine hydroxylase, the reported findings establish that Tyr179 is essential in maintaining the catalytic integrity and Phenylalanine binding of the enzyme via indirect interactions with the substrate, Phenylalanine. A model that accounts for the role of Tyr179 in binding Phenylalanine is proposed.
Csaba Paizs - One of the best experts on this subject based on the ideXlab platform.
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The production of l - and d -Phenylalanines using engineered Phenylalanine ammonia lyases from Petroselinum crispum
Scientific Reports, 2019Co-Authors: Souad Diana Tork, Emma Zsofia Aletta Nagy, Csaba Paizs, Monica Ioana Tosa, Lilla Cserepes, Diana Monica Bordea, Botond Nagy, László Csaba BenczeAbstract:The biocatalytic synthesis of l- and d-Phenylalanine analogues of high synthetic value have been developed using as biocatalysts mutant variants of Phenylalanine ammonia lyase from Petroselinum crispum (PcPAL), specifically tailored towards mono-substituted Phenylalanine and cinnamic acid substrates. The catalytic performance of the engineered PcPAL variants was optimized within the ammonia elimination and ammonia addition reactions, focusing on the effect of substrate concentration, biocatalyst:substrate ratio, reaction buffer and reaction time, on the conversion and enantiomeric excess values. The optimal conditions provided an efficient preparative scale biocatalytic procedure of valuable Phenylalanines, such as (S)-m-methoxyPhenylalanine (Y = 40%, ee > 99%), (S)-p-bromoPhenylalanine (Y = 82%, ee > 99%), (S)-m-(trifluoromethyl)Phenylalanine (Y = 26%, ee > 99%), (R)-p-methylPhenylalanine, (Y = 49%, ee = 95%) and (R)-m-(trifluoromethyl)Phenylalanine (Y = 34%, ee = 93%).
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mapping the hydrophobic substrate binding site of Phenylalanine ammonia lyase from petroselinum crispum
ACS Catalysis, 2019Co-Authors: Emma Zsofia Aletta Nagy, Souad Diana Tork, Pauline A Lang, Alina Filip, Florin Dan Irimie, Laszlo Poppe, Jurgen Brem, Christopher J. Schofield, Monica Ioana Tosa, Csaba PaizsAbstract:Modification of the hydrophobic binding pocket of Phenylalanine ammonia-lyase from Petroselinum crispum (PcPAL) enables increased activity and selectivity toward Phenylalanines and cinnamic acids m...
Jérôme Zoidakis - One of the best experts on this subject based on the ideXlab platform.
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Role of the second coordination sphere residue tyrosine 179 in substrate affinity and catalytic activity of Phenylalanine hydroxylase
JBIC Journal of Biological Inorganic Chemistry, 2004Co-Authors: Jérôme Zoidakis, Alon Volner, Kim Vu, Mahdi M. Abu-omarAbstract:Phenylalanine hydroxylase converts Phenylalanine to tyrosine utilizing molecular oxygen and tetrahydropterin as a cofactor, and belongs to the aromatic amino acid hydroxylases family. The catalytic domains of these enzymes are structurally similar. According to recent crystallographic studies, residue Tyr179 in Chromobacterium violaceum Phenylalanine hydroxylase is located in the active site and its hydroxyl oxygen is 5.1 Å from the iron, where it has been suggested to play a role in positioning the pterin cofactor. To determine the catalytic role of this residue, the point mutants Y179F and Y179A of Phenylalanine hydroxylase were prepared and characterized. Both mutants displayed comparable stability and metal binding to the native enzyme, as determined by their melting temperatures in the presence and absence of iron. The catalytic activity ( k _cat) of the Y179F and Y179A proteins was lower than wild-type Phenylalanine hydroxylase by an order of magnitude, suggesting that the hydroxyl group of Tyr179 plays a role in the rate-determining step in catalysis. The K _M values for different tetrahydropterin cofactors and Phenylalanine were decreased by a factor of 3–4 in the Y179F mutant. However, the K _M values for different pterin cofactors were slightly higher in the Y179A mutant than those measured for the wild-type enzyme, and, more significantly, the K _M value for Phenylalanine was increased by 10-fold in the Y179A mutant. By the criterion of k _cat/ K _Phe, the Y179F and Y179A mutants display 10% and 1%, respectively, of the activity of wild-type Phenylalanine hydroxylase. These results are consistent with Tyr179 having a pronounced role in binding Phenylalanine but a secondary effect in the formation of the hydroxylating species. In conjunction with recent crystallographic analyses of a ternary complex of Phenylalanine hydroxylase, the reported findings establish that Tyr179 is essential in maintaining the catalytic integrity and Phenylalanine binding of the enzyme via indirect interactions with the substrate, Phenylalanine. A model that accounts for the role of Tyr179 in binding Phenylalanine is proposed.
Paul F. Fitzpatrick - One of the best experts on this subject based on the ideXlab platform.
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Allosteric regulation of Phenylalanine hydroxylase
Archives of Biochemistry and Biophysics, 2011Co-Authors: Paul F. FitzpatrickAbstract:The liver enzyme Phenylalanine hydroxylase is responsible for conversion of excess Phenylalanine in the diet to tyrosine. Phenylalanine hydroxylase is activated by Phenylalanine; this activation is inhibited by the physiological reducing substrate tetrahydrobiopterin. Phosphorylation of Ser16 lowers the concentration of Phenylalanine for activation. This review discusses the present understanding of the molecular details of the allosteric regulation of the enzyme.
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Regulation of Phenylalanine hydroxylase
Biochemistry, 2010Co-Authors: Lawrence J. Dangott, Paul F. FitzpatrickAbstract:Phenylalanine acts as an allosteric activator of the tetrahydropterin-dependent enzyme Phenylalanine hydroxylase. Hydrogen/deuterium exchange monitored by mass spectrometry has been used to gain insight into local conformational changes accompanying activation of rat Phenylalanine hydroxylase by Phenylalanine. Peptides in the regulatory and catalytic domains that lie in the interface between these two domains show large increases in the extent of deuterium incorporation from solvent in the presence of Phenylalanine. In contrast, the effects of Phenylalanine on the exchange kinetics of a mutant enzyme lacking the regulatory domain are limited to peptides surrounding the binding site for the amino acid substrate. These results support a model in which the N-terminus of the protein acts as an inhibitory peptide, with Phenylalanine binding causing a conformational change in the regulatory domain that alters the interaction between the catalytic and regulatory domains.
Emma Zsofia Aletta Nagy - One of the best experts on this subject based on the ideXlab platform.
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The production of l - and d -Phenylalanines using engineered Phenylalanine ammonia lyases from Petroselinum crispum
Scientific Reports, 2019Co-Authors: Souad Diana Tork, Emma Zsofia Aletta Nagy, Csaba Paizs, Monica Ioana Tosa, Lilla Cserepes, Diana Monica Bordea, Botond Nagy, László Csaba BenczeAbstract:The biocatalytic synthesis of l- and d-Phenylalanine analogues of high synthetic value have been developed using as biocatalysts mutant variants of Phenylalanine ammonia lyase from Petroselinum crispum (PcPAL), specifically tailored towards mono-substituted Phenylalanine and cinnamic acid substrates. The catalytic performance of the engineered PcPAL variants was optimized within the ammonia elimination and ammonia addition reactions, focusing on the effect of substrate concentration, biocatalyst:substrate ratio, reaction buffer and reaction time, on the conversion and enantiomeric excess values. The optimal conditions provided an efficient preparative scale biocatalytic procedure of valuable Phenylalanines, such as (S)-m-methoxyPhenylalanine (Y = 40%, ee > 99%), (S)-p-bromoPhenylalanine (Y = 82%, ee > 99%), (S)-m-(trifluoromethyl)Phenylalanine (Y = 26%, ee > 99%), (R)-p-methylPhenylalanine, (Y = 49%, ee = 95%) and (R)-m-(trifluoromethyl)Phenylalanine (Y = 34%, ee = 93%).
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mapping the hydrophobic substrate binding site of Phenylalanine ammonia lyase from petroselinum crispum
ACS Catalysis, 2019Co-Authors: Emma Zsofia Aletta Nagy, Souad Diana Tork, Pauline A Lang, Alina Filip, Florin Dan Irimie, Laszlo Poppe, Jurgen Brem, Christopher J. Schofield, Monica Ioana Tosa, Csaba PaizsAbstract:Modification of the hydrophobic binding pocket of Phenylalanine ammonia-lyase from Petroselinum crispum (PcPAL) enables increased activity and selectivity toward Phenylalanines and cinnamic acids m...
