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Clive Harper - One of the best experts on this subject based on the ideXlab platform.
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Thiamine‐Dependent Enzyme Changes in the Brains of Alcoholics: Relationship to the Wernicke‐Korsakoff Syndrome
Alcoholism: Clinical and Experimental Research, 1993Co-Authors: Roger F. Butterworth, Jillian J Kril, Clive HarperAbstract:Chronic alcoholism results in thiamine deficiency as a consequence of poor nutrition, impaired absorption, and decreased phosphorylation to the Enzyme cofactor form of the vitamin, thiamine pyrophosphate (TPP). Results of this study demonstrate significant reductions of TPP-Dependent Enzymes [pyruvate dehydrogenase complex, α-ketoglutarate dehydrogenase (αKGDH), and transketolase] in autopsied cerebellar vermis samples from alcoholic patients with the clinical and neuropathologically confirmed diagnosis of Wernicke-Korsakoff Syndrome (WKS). Enzyme activities in brain samples from alcoholics without WKS were within normal limits and activities of a nonthiamine-Dependent Enzyme, glutamate dehydrogenase, were not significantly different from control values in brain samples from alcoholics with or without WKS. These findings provide evidence, for the first time, of a direct implication of TPP-related metabolic processes in the pathogenesis of WKS. Decreased activities of αKGDH could be the trigger for a sequence of metabolic events resulting in energy compromise, and ultimately neuronal death in this syndrome.
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thiamine Dependent Enzyme changes in the brains of alcoholics relationship to the wernicke korsakoff syndrome
Alcoholism: Clinical and Experimental Research, 1993Co-Authors: Roger F. Butterworth, Jillian J Kril, Clive HarperAbstract:Chronic alcoholism results in thiamine deficiency as a consequence of poor nutrition, impaired absorption, and decreased phosphorylation to the Enzyme cofactor form of the vitamin, thiamine pyrophosphate (TPP). Results of this study demonstrate significant reductions of TPP-Dependent Enzymes [pyruvate dehydrogenase complex, α-ketoglutarate dehydrogenase (αKGDH), and transketolase] in autopsied cerebellar vermis samples from alcoholic patients with the clinical and neuropathologically confirmed diagnosis of Wernicke-Korsakoff Syndrome (WKS). Enzyme activities in brain samples from alcoholics without WKS were within normal limits and activities of a nonthiamine-Dependent Enzyme, glutamate dehydrogenase, were not significantly different from control values in brain samples from alcoholics with or without WKS. These findings provide evidence, for the first time, of a direct implication of TPP-related metabolic processes in the pathogenesis of WKS. Decreased activities of αKGDH could be the trigger for a sequence of metabolic events resulting in energy compromise, and ultimately neuronal death in this syndrome.
Dominic J. Campopiano - One of the best experts on this subject based on the ideXlab platform.
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Research Article The Pyridoxal 5-Phosphate (PLP)-Dependent Enzyme Serine Palmitoyltransferase (SPT): Effects of the Small Subunits and Insights from Bacterial Mimics of Human hLCB2a
2016Co-Authors: Hsan Mutations, Jeffrey M. Harmon, Teresa M. Dunn, Dominic J. CampopianoAbstract:Copyright © 2013 Ashley E. Beattie et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The pyridoxal 5-phosphate (PLP)-Dependent Enzyme serine palmitoyltransferase (SPT) catalyses the first step of de novo sphingolipid biosynthesis. The core human Enzyme is a membrane-bound heterodimer composed of two subunits (hLCB1 and hLCB2a/b), and mutations in both hLCB1 (e.g., C133W and C133Y) and hLCB2a (e.g., V359M, G382V, and I504F) have been identified in patients with hereditary sensory and autonomic neuropathy type I (HSAN1), an inherited disorder that affects sensory and autonomic neurons. These mutations result in substrate promiscuity, leading to formation of neurotoxic deoxysphingolipids found in affected individuals. Here we measure the activities of the hLCB2a mutants in the presence of ssSPTa and ssSPTb and find that all decrease Enzyme activity. High resolution structural data of the homodimeric SPT Enzyme from the bacterium Sphingomonas paucimobilis (Sp SPT) provides a model to understand the impact of the hLCB2a mutations on the mechanism o
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the pyridoxal 5 phosphate plp Dependent Enzyme serine palmitoyltransferase spt effects of the small subunits and insights from bacterial mimics of human hlcb2a hsan1 mutations
BioMed Research International, 2013Co-Authors: Ashley E. Beattie, Jeffrey M. Harmon, Teresa M. Dunn, Sita D Gupta, Lenka Frankova, Agne Kazlauskaite, Dominic J. CampopianoAbstract:The pyridoxal 5′-phosphate (PLP)-Dependent Enzyme serine palmitoyltransferase (SPT) catalyses the first step of de novo sphingolipid biosynthesis. The core human Enzyme is a membrane-bound heterodimer composed of two subunits (hLCB1 and hLCB2a/b), and mutations in both hLCB1 (e.g., C133W and C133Y) and hLCB2a (e.g., V359M, G382V, and I504F) have been identified in patients with hereditary sensory and autonomic neuropathy type I (HSAN1), an inherited disorder that affects sensory and autonomic neurons. These mutations result in substrate promiscuity, leading to formation of neurotoxic deoxysphingolipids found in affected individuals. Here we measure the activities of the hLCB2a mutants in the presence of ssSPTa and ssSPTb and find that all decrease Enzyme activity. High resolution structural data of the homodimeric SPT Enzyme from the bacterium Sphingomonas paucimobilis (Sp SPT) provides a model to understand the impact of the hLCB2a mutations on the mechanism of SPT. The three human hLCB2a HSAN1 mutations map onto Sp SPT (V246M, G268V, and G385F), and these mutant mimics reveal that the amino acid changes have varying impacts; they perturb the PLP cofactor binding, reduce the affinity for both substrates, decrease the Enzyme activity, and, in the most severe case, cause the protein to be expressed in an insoluble form.
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Reconstitution of the pyridoxal 5'-phosphate (PLP) Dependent Enzyme serine palmitoyltransferase (SPT) with pyridoxal reveals a crucial role for the phosphate during catalysis
Chemical Communications, 2013Co-Authors: Ashley E. Beattie, John M. Wadsworth, David J. Clarke, Jonathan Lowther, Ho-lam Sin, Dominic J. CampopianoAbstract:The pyridoxal 5'-phosphate (PLP)-Dependent Enzyme serine palmitoyltransferase (SPT) is required for de novo sphingolipid biosynthesis. A previous study revealed a novel and unexpected interaction between the hydroxyl group of the l-serine substrate and the 5'-phosphate group of PLP. By using pyridoxal (PL), the dephosphorylated analogue of vitamin B6, we show here that this interaction is important for substrate specificity and optimal catalytic efficiency.
James H Naismith - One of the best experts on this subject based on the ideXlab platform.
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pmp diketopiperazine adducts form at the active site of a plp Dependent Enzyme involved in formycin biosynthesis
Chemical Communications, 2019Co-Authors: Sisi Gao, H Liu, Valerie De Crecylagard, Wen Zhu, Nigel G J Richards, James H NaismithAbstract:ForI is a PLP-Dependent Enzyme from the biosynthetic pathway of the C-nucleoside antibiotic formycin. Cycloserine is thought to inhibit PLP-Dependent Enzymes by irreversibly forming a PMP–isoxazole. We now report that ForI forms novel PMP–diketopiperazine derivatives following incubation with both D and L cycloserine. This unexpected result suggests chemical diversity in the chemistry of cycloserine inhibition.
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inhibition of the plp Dependent Enzyme serine palmitoyltransferase by cycloserine evidence for a novel decarboxylative mechanism of inactivation
Molecular BioSystems, 2010Co-Authors: Jonathan Lowther, David J. Clarke, Beverley A Yard, K A Johnson, L G Carter, Venugopal T Bhat, Marine C C Raman, Britta Ramakers, Stephen A Mcmahon, James H NaismithAbstract:Cycloserine (CS, 4-amino-3-isoxazolidone) is a cyclic amino acid mimic that is known to inhibit many essential pyridoxal 5′-phosphate (PLP)-Dependent Enzymes. Two CS enantiomers are known; D-cycloserine (DCS, also known as Seromycin) is a natural product that is used to treat resistant Mycobacterium tuberculosis infections as well as neurological disorders since it is a potent NMDA receptor agonist, and L-cycloserine (LCS) is a synthetic enantiomer whose usefulness as a drug has been hampered by its inherent toxicity arising through inhibition of sphingolipid metabolism. Previous studies on various PLP-Dependent Enzymes revealed a common mechanism of inhibition by both enantiomers of CS; the PLP cofactor is disabled by forming a stable 3-hydroxyisoxazole/pyridoxamine 5′-phosphate (PMP) adduct at the active site where the cycloserine ring remains intact. Here we describe a novel mechanism of CS inactivation of the PLP-Dependent Enzyme serine palmitoyltransferase (SPT) from Sphingomonas paucimobilis. SPT catalyses the condensation of L-serine and palmitoyl-CoA, the first step in the de novo sphingolipid biosynthetic pathway. We have used a range of kinetic, spectroscopic and structural techniques to postulate that both LCS and DCS inactivate SPT by transamination to form a free pyridoxamine 5′-phosphate (PMP) and β-aminooxyacetaldehyde that remain bound at the active site. We suggest this occurs by ring opening of the cycloserine ring followed by decarboxylation. Enzyme kinetics show that inhibition is reversed by incubation with excess PLP and that LCS is a more effective SPT inhibitor than DCS. UV-visible spectroscopic data, combined with site-directed mutagenesis, suggest that a mobile Arg378 residue is involved in cycloserine inactivation of SPT.
Roger F. Butterworth - One of the best experts on this subject based on the ideXlab platform.
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Thiamine‐Dependent Enzyme Changes in the Brains of Alcoholics: Relationship to the Wernicke‐Korsakoff Syndrome
Alcoholism: Clinical and Experimental Research, 1993Co-Authors: Roger F. Butterworth, Jillian J Kril, Clive HarperAbstract:Chronic alcoholism results in thiamine deficiency as a consequence of poor nutrition, impaired absorption, and decreased phosphorylation to the Enzyme cofactor form of the vitamin, thiamine pyrophosphate (TPP). Results of this study demonstrate significant reductions of TPP-Dependent Enzymes [pyruvate dehydrogenase complex, α-ketoglutarate dehydrogenase (αKGDH), and transketolase] in autopsied cerebellar vermis samples from alcoholic patients with the clinical and neuropathologically confirmed diagnosis of Wernicke-Korsakoff Syndrome (WKS). Enzyme activities in brain samples from alcoholics without WKS were within normal limits and activities of a nonthiamine-Dependent Enzyme, glutamate dehydrogenase, were not significantly different from control values in brain samples from alcoholics with or without WKS. These findings provide evidence, for the first time, of a direct implication of TPP-related metabolic processes in the pathogenesis of WKS. Decreased activities of αKGDH could be the trigger for a sequence of metabolic events resulting in energy compromise, and ultimately neuronal death in this syndrome.
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thiamine Dependent Enzyme changes in the brains of alcoholics relationship to the wernicke korsakoff syndrome
Alcoholism: Clinical and Experimental Research, 1993Co-Authors: Roger F. Butterworth, Jillian J Kril, Clive HarperAbstract:Chronic alcoholism results in thiamine deficiency as a consequence of poor nutrition, impaired absorption, and decreased phosphorylation to the Enzyme cofactor form of the vitamin, thiamine pyrophosphate (TPP). Results of this study demonstrate significant reductions of TPP-Dependent Enzymes [pyruvate dehydrogenase complex, α-ketoglutarate dehydrogenase (αKGDH), and transketolase] in autopsied cerebellar vermis samples from alcoholic patients with the clinical and neuropathologically confirmed diagnosis of Wernicke-Korsakoff Syndrome (WKS). Enzyme activities in brain samples from alcoholics without WKS were within normal limits and activities of a nonthiamine-Dependent Enzyme, glutamate dehydrogenase, were not significantly different from control values in brain samples from alcoholics with or without WKS. These findings provide evidence, for the first time, of a direct implication of TPP-related metabolic processes in the pathogenesis of WKS. Decreased activities of αKGDH could be the trigger for a sequence of metabolic events resulting in energy compromise, and ultimately neuronal death in this syndrome.
Masao Ohashi - One of the best experts on this subject based on the ideXlab platform.
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sam Dependent Enzyme catalysed pericyclic reactions in natural product biosynthesis
Nature, 2017Co-Authors: Masao Ohashi, Mengbin Chen, Mancheng Tang, Zhongyue Yang, Michio Sato, Kenji Watanabe, K N Houk, Yi TangAbstract:The Enzyme LepI is found to be capable of catalysing several natural-product pericyclic transformations, including a hetero-Diels–Alder reaction and a retro-Claisen rearrangement. Although common in synthesis, naturally occurring pericyclic reactions, in which two fragments combine to form a cyclic molecule, are rare. Several examples of cyclohexene-forming Enzymes called Diels–Alderases have been discovered. However, biosynthetic inverse electron demand Diels–Alder reactions are still unknown. These reactions often involve heteroatoms (non-carbon atoms) in the cycloaddition step, so are important in the synthesis of both heterocyclic and natural products. Here, the authors report the versatile S-adenosyl-L-methionine (SAM)-Dependent Enzyme, LepI, which is capable of catalysing several pericyclic transformations, including a hetero-Diels–Alder reaction. The biosynthesis of the cytotoxic leporin B proceeds via a bifurcated reaction pathway regulated by LepI, a direct hetero-Diels–Alder reaction and an indirect Diels–Alder/retro-Claisen rearrangement sequence, converging to give the heterocyclic pyran product. Pericyclic reactions—which proceed in a concerted fashion through a cyclic transition state—are among the most powerful synthetic transformations used to make multiple regioselective and stereoselective carbon–carbon bonds1. They have been widely applied to the synthesis of biologically active complex natural products containing contiguous stereogenic carbon centres2,3,4,5,6. Despite the prominence of pericyclic reactions in total synthesis, only three naturally existing enzymatic examples (the intramolecular Diels–Alder reaction7, and the Cope8 and the Claisen rearrangements9) have been characterized. Here we report a versatile S-adenosyl-l-methionine (SAM)-Dependent Enzyme, LepI, that can catalyse stereoselective dehydration followed by three pericyclic transformations: intramolecular Diels–Alder and hetero-Diels–Alder reactions via a single ambimodal transition state, and a retro-Claisen rearrangement. Together, these transformations lead to the formation of the dihydropyran core of the fungal natural product, leporin10. Combined in vitro enzymatic characterization and computational studies provide insight into how LepI regulates these bifurcating biosynthetic reaction pathways by using SAM as the cofactor. These pathways converge to the desired biosynthetic end product via the (SAM-Dependent) retro-Claisen rearrangement catalysed by LepI. We expect that more pericyclic biosynthetic enzymatic transformations remain to be discovered in naturally occurring Enzyme ‘toolboxes’11. The new role of the versatile cofactor SAM is likely to be found in other examples of Enzyme catalysis.
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sam Dependent Enzyme catalysed pericyclic reactions in natural product biosynthesis
Nature, 2017Co-Authors: Masao Ohashi, Mengbin Chen, Mancheng Tang, Zhongyue Yang, Michio Sato, Fang Liu, Yang Hai, Kenji WatanabeAbstract:Pericyclic reactions-which proceed in a concerted fashion through a cyclic transition state-are among the most powerful synthetic transformations used to make multiple regioselective and stereoselective carbon-carbon bonds. They have been widely applied to the synthesis of biologically active complex natural products containing contiguous stereogenic carbon centres. Despite the prominence of pericyclic reactions in total synthesis, only three naturally existing enzymatic examples (the intramolecular Diels-Alder reaction, and the Cope and the Claisen rearrangements) have been characterized. Here we report a versatile S-adenosyl-l-methionine (SAM)-Dependent Enzyme, LepI, that can catalyse stereoselective dehydration followed by three pericyclic transformations: intramolecular Diels-Alder and hetero-Diels-Alder reactions via a single ambimodal transition state, and a retro-Claisen rearrangement. Together, these transformations lead to the formation of the dihydropyran core of the fungal natural product, leporin. Combined in vitro enzymatic characterization and computational studies provide insight into how LepI regulates these bifurcating biosynthetic reaction pathways by using SAM as the cofactor. These pathways converge to the desired biosynthetic end product via the (SAM-Dependent) retro-Claisen rearrangement catalysed by LepI. We expect that more pericyclic biosynthetic enzymatic transformations remain to be discovered in naturally occurring Enzyme 'toolboxes'. The new role of the versatile cofactor SAM is likely to be found in other examples of Enzyme catalysis.
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sam Dependent Enzyme catalysed pericyclic reactions in natural product biosynthesis
Nature, 2017Co-Authors: Masao Ohashi, Mengbin Chen, Mancheng Tang, Zhongyue Yang, Michio Sato, Fang Liu, Yang Hai, Kenji WatanabeAbstract:The Enzyme LepI is found to be capable of catalysing several natural-product pericyclic transformations, including a hetero-Diels–Alder reaction and a retro-Claisen rearrangement.