Regulation of an important glycolytic enzyme, pyruvate kinase, through phosphorylation in the larvae of a species of freeze‐tolerant insect, Eurosta solidaginis

2020 ◽  
Author(s):  
J. Abboud ◽  
S. R. Green ◽  
M. B. Smolinski ◽  
K. B. Storey
Keyword(s):  
Pyruvate Kinase ◽  
Glycolytic Enzyme ◽  
Eurosta Solidaginis ◽  
Freeze Tolerant

scholarly journals Changes in activities and isozyme patterns of glycolytic enzymes during erythroid differentiation in vitro

Blood ◽  
1984 ◽  
Vol 64 (3) ◽  
pp. 607-613 ◽  
Author(s):  
W Nijhof ◽  
PK Wierenga ◽  
GE Staal ◽  
G Jansen
Keyword(s):  
Pyruvate Kinase ◽  
Gradient Centrifugation ◽  
Recovery Period ◽  
Erythroid Differentiation ◽  
Glycolytic Enzyme ◽  
Minor Amount ◽  
Type I ◽  
In Vitro Differentiation ◽  
Percoll Density Gradient Centrifugation

Late committed progenitor cells of erythropoiesis, CFU-E (colony- forming unit--erythroid), were isolated from mouse spleens to near homogeneity by a three-step enrichment procedure. The procedure included a four-day pretreatment of bled mice with the antibiotic thiamphenicol, a recovery period of 3 1/2 days, followed by centrifugal elutriation and Percoll density gradient centrifugation of the spleen cells. This practically pure CFU-E population was used to study some aspects of erythroid differentiation in vitro. Colony growth, as well as morphology and glycolytic enzyme activities of cells isolated at selected times of the 48-hour culture period, were determined. Marked declining activities of several enzymes, including hexokinase, phosphofructokinase, aldolase, enolase, pyruvate kinase, and glucose-6- phosphate dehydrogenase, were observed during in vitro differentiation. The activity of diphosphoglycerate mutase was almost absent in the CFU- E, but progressively increased during differentiation. The isozyme distribution of aldolase and enolase did not change during CFU-E in vitro differentiation into the reticulocyte. Hexokinase (HK) in the CFU- E contained mainly a double-banded type I isozyme, in addition to a minor amount of HK II. During differentiation, a shift was noticed within the double-banded HK I region, whereas HK ii disappeared after one cell division. Pyruvate kinase in the CFU-E was characterized by the presence of both the K-type and the L-type isozyme and hybrids of these isozyme types. During in vitro differentiation, the production of the K-type isozyme rapidly stops in favor of the L type.

scholarly journals Functional cross-talk between allosteric effects of activating and inhibiting ligands underlies PKM2 regulation

2018 ◽  
Author(s):  
Jamie A. Macpherson ◽  
Alina Theisen ◽  
Laura Masino ◽  
Louise Fets ◽  
Paul C. Driscoll ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
High Activity ◽  
Pyruvate Kinase ◽  
Cross Talk ◽  
Glycolytic Enzyme ◽  
Computational Framework ◽  
Allosteric Inhibitor ◽  
Fructose 1,6 Bisphosphate ◽  
Low Activity

ABSTRACTAllosteric regulation is central to the role of the glycolytic enzyme pyruvate kinase M2 (PKM2) in cellular metabolism. Multiple activating and inhibitory allosteric ligands regulate PKM2 activity by controlling the equilibrium between high activity tetramers and low activity dimers and monomers. However, it remains elusive how allosteric inputs upon simultaneous binding of different ligands are integrated to regulate PKM2 activity. Here, we show that, in the presence of the allosteric inhibitor L-phenylalanine (Phe), the activator fructose 1,6-bisphosphate (FBP) can induce PKM2 tetramerisation, but fails to maximally increase enzymatic activity. Guided by a new computational framework we developed to identify residues that mediate FBP-induced allostery, we generated two PKM2 mutants, A327S and C358A, in which activation by FBP remains intact but cannot be attenuated by Phe. Our findings demonstrate a role for residues involved in FBP-induced allostery in enabling the integration of allosteric input from Phe and reveal a mechanism that underlies the co-ordinate regulation of PKM2 activity by multiple allosteric ligands.

scholarly journals Pyruvate Kinase (PK) Deficiency Hereditary Nonspherocytic Hemolytic Anemia

Blood ◽  
1962 ◽  
Vol 19 (3) ◽  
pp. 267-295 ◽  
Author(s):  
KOUICHI R. TANAKA ◽  
WILLIAM N. VALENTINE ◽  
SHIRO MIWA
Keyword(s):  
Pyruvate Kinase ◽  
Hemolytic Anemia ◽  
Glycolytic Enzyme ◽  
Enzyme Deficiency ◽  
Exact Nature ◽  
Symptomatic Disease ◽  
Genetically Determined ◽  
Recessive Transmission ◽  
Enzymatic Defect

Abstract 1. The erythrocytes of seven patients conforming to the criteria of Type II congenital nonspherocytic hemolytic anemia have been demonstrated to have a specific deficiency in the glycolytic enzyme pyruvate kinase. Other glycolytic enzymes, glucose-6-phosphate and 6-phosphogluconic dehydrogenases, and certain non-glycolytic erythrocyte enzymes are normally active. The leukocytes of these patients possess normal pyruvate kinase activity. 2. Although no inhibitors were detected, the exact nature of the enzymatic defect remains to be elucidated. 3. Family studies provide strong evidence for a genetically determined disorder and are consistent with an autosomal recessive transmission of the defect. A partial enzyme deficiency, not reflected in clinical disease, is present in heterozygotes. The symptomatic disease, though variable in severity, appears to be due to homozygosity for the defect. 4. It is suggested that the enzyme deficiency is pathogenetically related to the premature demise of the red cells in vivo. 5. The name "pyruvate kinase (PK) deficiency hereditary nonspherocytic hemolytic anemia" is proposed for these patients.

scholarly journals Biochemical and structural insights into how amino acids regulate pyruvate kinase muscle isoform 2

2020 ◽  
Vol 295 (16) ◽  
pp. 5390-5403 ◽  
Author(s):  
Suparno Nandi ◽  
Mishtu Dey
Keyword(s):  
Pyruvate Kinase ◽  
Cancer Metabolism ◽  
Gel Filtration ◽  
Binding Pocket ◽  
Glycolytic Enzyme ◽  
X Ray Crystallography ◽  
Highly Active ◽  
Attractive Therapeutic Target ◽  
Fluorescence Binding ◽  
Small Molecule Modulators

Pyruvate kinase muscle isoform 2 (PKM2) is a key glycolytic enzyme involved in ATP generation and critical for cancer metabolism. PKM2 is expressed in many human cancers and is regulated by complex mechanisms that promote tumor growth and proliferation. Therefore, it is considered an attractive therapeutic target for modulating tumor metabolism. Various stimuli allosterically regulate PKM2 by cycling it between highly active and less active states. Several small molecules activate PKM2 by binding to its intersubunit interface. Serine and cysteine serve as an activator and inhibitor of PKM2, respectively, by binding to its amino acid (AA)-binding pocket, which therefore represents a potential druggable site. Despite binding similarly to PKM2, how cysteine and serine differentially regulate this enzyme remains elusive. Using kinetic analyses, fluorescence binding, X-ray crystallography, and gel filtration experiments with asparagine, aspartate, and valine as PKM2 ligands, we examined whether the differences in the side-chain polarity of these AAs trigger distinct allosteric responses in PKM2. We found that Asn (polar) and Asp (charged) activate PKM2 and that Val (hydrophobic) inhibits it. The results also indicate that both Asn and Asp can restore the activity of Val-inhibited PKM2. AA-bound crystal structures of PKM2 displayed distinctive interactions within the binding pocket, causing unique allosteric effects in the enzyme. These structure-function analyses of AA-mediated PKM2 regulation shed light on the chemical requirements in the development of mechanism-based small-molecule modulators targeting the AA-binding pocket of PKM2 and provide broader insights into the regulatory mechanisms of complex allosteric enzymes.

scholarly journals Changes in activities and isozyme patterns of glycolytic enzymes during erythroid differentiation in vitro

Blood ◽  
1984 ◽  
Vol 64 (3) ◽  
pp. 607-613 ◽  
Author(s):  
W Nijhof ◽  
PK Wierenga ◽  
GE Staal ◽  
G Jansen
Keyword(s):  
Pyruvate Kinase ◽  
Gradient Centrifugation ◽  
Recovery Period ◽  
Erythroid Differentiation ◽  
Glycolytic Enzyme ◽  
Minor Amount ◽  
Type I ◽  
In Vitro Differentiation ◽  
Percoll Density Gradient Centrifugation

Abstract Late committed progenitor cells of erythropoiesis, CFU-E (colony- forming unit--erythroid), were isolated from mouse spleens to near homogeneity by a three-step enrichment procedure. The procedure included a four-day pretreatment of bled mice with the antibiotic thiamphenicol, a recovery period of 3 1/2 days, followed by centrifugal elutriation and Percoll density gradient centrifugation of the spleen cells. This practically pure CFU-E population was used to study some aspects of erythroid differentiation in vitro. Colony growth, as well as morphology and glycolytic enzyme activities of cells isolated at selected times of the 48-hour culture period, were determined. Marked declining activities of several enzymes, including hexokinase, phosphofructokinase, aldolase, enolase, pyruvate kinase, and glucose-6- phosphate dehydrogenase, were observed during in vitro differentiation. The activity of diphosphoglycerate mutase was almost absent in the CFU- E, but progressively increased during differentiation. The isozyme distribution of aldolase and enolase did not change during CFU-E in vitro differentiation into the reticulocyte. Hexokinase (HK) in the CFU- E contained mainly a double-banded type I isozyme, in addition to a minor amount of HK II. During differentiation, a shift was noticed within the double-banded HK I region, whereas HK ii disappeared after one cell division. Pyruvate kinase in the CFU-E was characterized by the presence of both the K-type and the L-type isozyme and hybrids of these isozyme types. During in vitro differentiation, the production of the K-type isozyme rapidly stops in favor of the L type.

Regulation of pyruvate kinase from muscle of Freeze tolerant wood frog, Rana sylvatica

Cryobiology ◽  
2016 ◽  
Vol 73 (3) ◽  
pp. 437
Author(s):  
J. Mattice ◽  
M. Smolinski ◽  
K. Storey
Keyword(s):  
Pyruvate Kinase ◽  
Rana Sylvatica ◽  
Wood Frog ◽  
Freeze Tolerant

Red Cell Pyruvate Kinase Deficiency: Molecular Characterization of 10 New Variants.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1590-1590
Author(s):  
Elisa Fermo ◽  
Paola Bianchi ◽  
Cristina Vercellati ◽  
Frederic Cotton ◽  
Alberto Zanella
Keyword(s):  
Pyruvate Kinase ◽  
Direct Sequencing ◽  
Large Deletion ◽  
Glycolytic Enzyme ◽  
Coding Region ◽  
Clinical Pattern ◽  
Pyruvate Kinase Deficiency ◽  
Exon 11 ◽  
Flanking Regions

Abstract PK deficiency is the most common glycolytic enzyme defect associated with chronic non-spherocytic hemolytic anemia. To date about 150 different mutations have been identified in the PK-LR gene. Among them only one large deletion has been described in Gipsy resulting in the loss of exon 11. We report 10 new variants of LR-PK gene in 8 families with pyruvate kinase deficiency. The entire coding region and intronic flanking regions were analyzed by direct sequencing. The results of the molecular analysis are reported in the table: Pt Origin Hb (g/dL) Tx (n.) PK Activity (IU/gHb) Mutation Aminoacidic substitution SD Italy 15.1 0 10.6 107G / ? Ala36Gly/? TT Australia 8.9 0 nd 409A / del5006bp( IVS3-nt 1431) Ala137Thr / del ex 4-11 NR Italy 9 >50 5.5 661A /1209A Asp221Asn /Met403Ile NA Italy 9 >50 5.3 661A /1209A Asp221Asn /Met403Ile SA Italy 12 0 5.5 1456T/ 1209A Arg486Trp/ Met403Ile SC Italy nd nd 5.6 1529A/ 859C Arg510Gln/ Phe287Leu CM Italy 9.5 0 13.6 1456T/ 958A Arg486Trp/ Val320Met PS Italy 13.2 0 8 1094T /? Lys365Met /? VR Italy 10.5 0 10.7 1706A / ? Arg569Gln /? GR Guinea 14.4 0 7.6 1269A/ IVS9+43c Splice site/? Ref. Values 12.2–16 11.1–15.5 Mutations reported in bold are new. By comparing the amino acids sequences among several species (cat M1, chicken M, rat L, yeast and human), we found that mutations 661A, 859C, 958A, 1094T and 1209A involve highly conserved residues. Mutation 1209A when present in association with 661A (cases NA and NR) results in a severe clinical pattern with need of transfusion support, whereas in compound heterozygosity with 1456T (case SA, mother of NA and NR) is associated with a less severe clinical pattern. The variant 1706A was found in a patient carrying the polymorphism 1705C at the homozygous level; the mutation in association with the polymorphism determines the aminoacidic substitution Arg596Gln. A deletion of 5006 nucleotides extending from intron 3 to the last 3 nucleotides of exon 10 has been found in an Australian baby dead at birth (case TT); the mutation results in a large cDNA deletion encompassing exon 4 and exon11 included. This is the largest abnormality so far detected in LR-PK gene.

scholarly journals Glycolytic enzyme expression and pyruvate kinase activity in cultured fibroblasts from type 1 diabetic patients with and without nephropathy

2008 ◽  
Vol 1782 (11) ◽  
pp. 627-633 ◽  
Author(s):  
Elisabetta Iori ◽  
Renato Millioni ◽  
Lucia Puricelli ◽  
Giorgio Arrigoni ◽  
Livia Lenzini ◽  
...  
Keyword(s):  
Pyruvate Kinase ◽  
Kinase Activity ◽  
Glycolytic Enzyme ◽  
Diabetic Patients ◽  
Enzyme Expression ◽  
Pyruvate Kinase Activity ◽  
Cultured Fibroblasts ◽  
Type 1 Diabetic Patients

scholarly journals Insulin enhances metabolic capacities of cancer cells by dual regulation of glycolytic enzyme pyruvate kinase M2

2013 ◽  
Vol 12 (1) ◽  
pp. 72 ◽  
Author(s):  
Mohd Iqbal ◽  
Farid Siddiqui ◽  
Vibhor Gupta ◽  
Shilpi Chattopadhyay ◽  
Prakasam Gopinath ◽  
...  
Keyword(s):  
Cancer Cells ◽  
Pyruvate Kinase ◽  
Glycolytic Enzyme ◽  
Pyruvate Kinase M2
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