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Kleczkowski, Leszek A.ORCID iD iconorcid.org/0000-0001-8685-9665
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Publications (10 of 75) Show all publications
Kleczkowski, L. A. & Igamberdiev, A. U. (2024). Multiple roles of glycerate kinase—from photorespiration to gluconeogenesis, C4 metabolism, and plant immunity. International Journal of Molecular Sciences, 25(6), Article ID 3258.
Open this publication in new window or tab >>Multiple roles of glycerate kinase—from photorespiration to gluconeogenesis, C4 metabolism, and plant immunity
2024 (English)In: International Journal of Molecular Sciences, ISSN 1661-6596, E-ISSN 1422-0067, Vol. 25, no 6, article id 3258Article, review/survey (Refereed) Published
Abstract [en]

Plant glycerate kinase (GK) was previously considered an exclusively chloroplastic enzyme of the glycolate pathway (photorespiration), and its sole predicted role was to return most of the glycolate-derived carbon (as glycerate) to the Calvin cycle. However, recent discovery of cytosolic GK revealed metabolic links for glycerate to other processes. Although GK was initially proposed as being solely regulated by substrate availability, subsequent discoveries of its redox regulation and the light involvement in the production of chloroplastic and cytosolic GK isoforms have indicated a more refined regulation of the pathways of glycerate conversion. Here, we re-evaluate the importance of GK and emphasize its multifaceted role in plants. Thus, GK can be a major player in several branches of primary metabolism, including the glycolate pathway, gluconeogenesis, glycolysis, and C4 metabolism. In addition, recently, the chloroplastic (but not cytosolic) GK isoform was implicated as part of a light-dependent plant immune response to pathogen attack. The origins of glycerate are also discussed here; it is produced in several cell compartments and undergoes huge fluctuations depending on light/dark conditions. The recent discovery of the vacuolar glycerate transporter adds yet another layer to our understanding of glycerate transport/metabolism and that of other two- and three-carbon metabolites.

Place, publisher, year, edition, pages
MDPI, 2024
Keywords
C4 photosynthesis, gluconeogenesis, glycerate metabolism, glycolate pathway, Phytophthora infestans, sucrose synthesis
National Category
Botany
Identifiers
urn:nbn:se:umu:diva-223078 (URN)10.3390/ijms25063258 (DOI)001193343000001 ()38542231 (PubMedID)2-s2.0-85189108088 (Scopus ID)
Funder
Umeå University
Available from: 2024-04-15 Created: 2024-04-15 Last updated: 2024-04-15Bibliographically approved
Decker, D., Aubert, J., Wilczynska, M. & Kleczkowski, L. A. (2023). Exploring redox modulation of plant UDP-glucose pyrophosphorylase. International Journal of Molecular Sciences, 24(10), Article ID 8914.
Open this publication in new window or tab >>Exploring redox modulation of plant UDP-glucose pyrophosphorylase
2023 (English)In: International Journal of Molecular Sciences, ISSN 1661-6596, E-ISSN 1422-0067, Vol. 24, no 10, article id 8914Article in journal (Refereed) Published
Abstract [en]

UDP-glucose (UDPG) pyrophosphorylase (UGPase) catalyzes a reversible reaction, producing UDPG, which serves as an essential precursor for hundreds of glycosyltransferases in all organisms. In this study, activities of purified UGPases from sugarcane and barley were found to be reversibly redox modulated in vitro through oxidation by hydrogen peroxide or oxidized glutathione (GSSG) and through reduction by dithiothreitol or glutathione. Generally, while oxidative treatment decreased UGPase activity, a subsequent reduction restored the activity. The oxidized enzyme had increased Km values with substrates, especially pyrophosphate. The increased Km values were also observed, regardless of redox status, for UGPase cysteine mutants (Cys102Ser and Cys99Ser for sugarcane and barley UGPases, respectively). However, activities and substrate affinities (Kms) of sugarcane Cys102Ser mutant, but not barley Cys99Ser, were still prone to redox modulation. The data suggest that plant UGPase is subject to redox control primarily via changes in the redox status of a single cysteine. Other cysteines may also, to some extent, contribute to UGPase redox status, as seen for sugarcane enzymes. The results are discussed with respect to earlier reported details of redox modulation of eukaryotic UGPases and regarding the structure/function properties of these proteins.

Place, publisher, year, edition, pages
MDPI, 2023
Keywords
carbohydrate metabolism, glutathione, hydrogen peroxide, protein structure, redox regulation, substrate affinity, UDP-glucose pyrophosphorylase
National Category
Biochemistry and Molecular Biology Botany
Identifiers
urn:nbn:se:umu:diva-209182 (URN)10.3390/ijms24108914 (DOI)000997890000001 ()37240260 (PubMedID)2-s2.0-85160372737 (Scopus ID)
Funder
The Kempe FoundationsLars Hierta Memorial Foundation
Available from: 2023-06-20 Created: 2023-06-20 Last updated: 2023-06-20Bibliographically approved
Kleczkowski, L. A. & Igamberdiev, A. U. (2023). Magnesium and cell energetics: at the junction of metabolism of adenylate and non-adenylate nucleotides. Journal of plant physiology (Print), 280, Article ID 153901.
Open this publication in new window or tab >>Magnesium and cell energetics: at the junction of metabolism of adenylate and non-adenylate nucleotides
2023 (English)In: Journal of plant physiology (Print), ISSN 0176-1617, E-ISSN 1618-1328, Vol. 280, article id 153901Article in journal (Refereed) Published
Abstract [en]

Free magnesium (Mg2+) represents a powerful signal arising from interconversions of adenylates (ATP, ADP and AMP). This is a consequence of the involvement of adenylate kinase (AK) which equilibrates adenylates and uses defined species of Mg-complexed and Mg-free adenylates in both directions of its reaction. However, cells contain also other reversible Mg2+-dependent enzymes that equilibrate non-adenylate nucleotides (uridylates, cytidylates and guanylates), i.e. nucleoside monophosphate kinases (NMPKs) and nucleoside diphosphate kinase (NDPK). Here, we propose that AK activity is tightly coupled to activities of NMPK and NDPK, linking adenylate equilibrium to equilibria of other nucleotides, and with [Mg2+] controlling the ratios of Mg-chelated and Mg-free nucleotides. This coupling establishes main hubs for adenylate-driven equilibration of non-adenylate nucleotides, with [Mg2+] acting as signal arising from all nucleotides rather than adenylates only. Further consequences involve an overall adenylate control of UTP-, GTP- and CTP-dependent pathways and the availability of substrates for RNA and DNA synthesis.

Place, publisher, year, edition, pages
Elsevier, 2023
Keywords
Adenylate kinase, Guanylate kinase, Magnesium signaling, Nucleoside diphosphate kinase, Nucleoside monophosphate kinase, Uridylate-cytidylate kinase
National Category
Botany Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-202227 (URN)10.1016/j.jplph.2022.153901 (DOI)000911805900001 ()36549033 (PubMedID)2-s2.0-85144811958 (Scopus ID)
Available from: 2023-01-10 Created: 2023-01-10 Last updated: 2023-09-05Bibliographically approved
Igamberdiev, A. U. & Kleczkowski, L. A. (2023). Toward understanding the emergence of life: a dual function of the system of nucleotides in the metabolically closed autopoietic organization. Biosystems (Amsterdam. Print), 224, Article ID 104837.
Open this publication in new window or tab >>Toward understanding the emergence of life: a dual function of the system of nucleotides in the metabolically closed autopoietic organization
2023 (English)In: Biosystems (Amsterdam. Print), ISSN 0303-2647, E-ISSN 1872-8324, Vol. 224, article id 104837Article in journal (Refereed) Published
Abstract [en]

General structure of metabolism includes the reproduction of catalysts that govern metabolism. In this structure, the system becomes autopoietic in the sense of Maturana and Varela, and it is closed to efficient causation as defined by Robert Rosen. The autopoietic maintenance and operation of the catalysts takes place via the set of free nucleotides while the synthesis of catalysts occurs via the information encoded by the set of nucleotides arranged in polymers of RNA and DNA. Both energy charge and genetic information use the components of the same pool of nucleoside triphosphates, which is equilibrated by thermodynamic buffering enzymes such as nucleoside diphosphate kinase and adenylate kinase. This occurs in a way that the system becomes internally stable and metabolically closed, which initially could be realized at the level of ribozymes catalyzing basic metabolic reactions as well as own reproduction. The function of ATP, GTP, UTP, and CTP is dual, as these species participate both in the general metabolism as free nucleotides and in the transfer of genetic information via covalent polymerization to nucleic acids. The changes in their pools directly impact both bioenergetic pathways and nucleic acid turnover. Here we outline the concept of metabolic closure of biosystems grounded in the dual function of nucleotide coenzymes that serve both as energetic and informational molecules and through this duality generate the autopoietic performance and the ability for codepoietic evolutionary transformations of living systems starting from the emergence of prebiotic systems.

Place, publisher, year, edition, pages
Elsevier, 2023
Keywords
Autopoiesis, Codepoiesis, Coenzyme, Metabolic closure, Nucleoside triphosphates, Ribozymes, Thermodynamic buffering
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-204078 (URN)10.1016/j.biosystems.2023.104837 (DOI)000960802100001 ()2-s2.0-85146310966 (Scopus ID)
Funder
Umeå University
Available from: 2023-01-27 Created: 2023-01-27 Last updated: 2023-09-05Bibliographically approved
Kleczkowski, L. A. & Decker, D. (2022). Effects of Magnesium, Pyrophosphate and Phosphonates on Pyrophosphorolytic Reaction of UDP-Glucose Pyrophosphorylase. PLANTS, 11(12), Article ID 1611.
Open this publication in new window or tab >>Effects of Magnesium, Pyrophosphate and Phosphonates on Pyrophosphorolytic Reaction of UDP-Glucose Pyrophosphorylase
2022 (English)In: PLANTS, E-ISSN 2223-7747, Vol. 11, no 12, article id 1611Article in journal (Refereed) Published
Abstract [en]

UDP-glucose pyrophosphorylase (UGPase) carries a freely reversible reaction, using glucose-1-P and UTP to produce UDP-glucose (UDPG) and pyrophosphate (PPi ), with UDPG being essential for glycosylation reactions in all organisms including, e.g., synthesis of sucrose, cellulose and glycoproteins. In the present study, we found that free magnesium (Mg2+) had profound effects on the reverse reaction of purified barley UGPase, and was absolutely required for its activity, with an apparent Km of 0.13 mM. More detailed analyses with varied concentrations of MgPPi allowed us to conclude that it is the MgPPi complex which serves as true substrate for UGPase in its reverse reaction, with an apparent Km of 0.06 mM. Free PPi was an inhibitor in this reaction. Given the key role of PPi in the UGPase reaction, we have also tested possible effects of phosphonates, which are analogs of PPi and phosphate (Pi ). Clodronate and etidronate (PPi analogs) had little or no effect on UGPase activity, whereas fosetyl-Al (Pi analog), a known fungicide, acted as effective near-competitive inhibitor versus PPi, with Ki of 0.15 mM. The data are discussed with respect to the role of magnesium in the UGPase reaction and elucidating the use of inhibitors in studies on cellular function of UGPase and related enzymes.

Place, publisher, year, edition, pages
MDPI, 2022
Keywords
chemical genetics, Dixon plot, fosetyl-Al, inhibitor kinetics, magnesium activation, Phytophthora
National Category
Botany
Identifiers
urn:nbn:se:umu:diva-203179 (URN)10.3390/plants11121611 (DOI)000816007800001 ()35736762 (PubMedID)2-s2.0-85132156979 (Scopus ID)
Funder
Lars Hierta Memorial FoundationUmeå University
Available from: 2023-01-16 Created: 2023-01-16 Last updated: 2023-01-16Bibliographically approved
Kleczkowski, L. A. & Igamberdiev, A. U. (2021). Magnesium signaling in plants. International Journal of Molecular Sciences, 22(3), Article ID 1159.
Open this publication in new window or tab >>Magnesium signaling in plants
2021 (English)In: International Journal of Molecular Sciences, ISSN 1661-6596, E-ISSN 1422-0067, Vol. 22, no 3, article id 1159Article, review/survey (Refereed) Published
Abstract [en]

Free magnesium (Mg2+) is a signal of the adenylate (ATP+ADP+AMP) status in the cells. It results from the equilibrium of adenylate kinase (AK), which uses Mg-chelated and Mgfree adenylates as substrates in both directions of its reaction. The AK-mediated primary control of intracellular [Mg2+] is finely interwoven with the operation of membrane-bound adenylateand Mg2+-translocators, which in a given compartment control the supply of free adenylates and Mg2+ for the AK-mediated equilibration. As a result, [Mg2+] itself varies both between and within the compartments, depending on their energetic status and environmental clues. Other key nucleotide-utilizing/producing enzymes (e.g., nucleoside diphosphate kinase) may also be involved in fine-tuning of the intracellular [Mg2+]. Changes in [Mg2+] regulate activities of myriads of Mgutilizing/ requiring enzymes, affecting metabolism under both normal and stress conditions, and impacting photosynthetic performance, respiration, phloem loading and other processes. In compartments controlled by AK equilibrium (cytosol, chloroplasts, mitochondria, nucleus), the intracellular [Mg2+] can be calculated from total adenylate contents, based on the dependence of the apparent equilibrium constant of AK on [Mg2+]. Magnesium signaling, reflecting cellular adenylate status, is likely widespread in all eukaryotic and prokaryotic organisms, due simply to the omnipresent nature of AK and to its involvement in adenylate equilibration.

Place, publisher, year, edition, pages
MPDI, 2021
Keywords
Adenylate energy charge, Adenylate kinase, Cellular magnesium, Free magnesium, Nucleoside diphosphate kinase, Thermodynamic buffering
National Category
Botany Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-180547 (URN)10.3390/ijms22031159 (DOI)000615312800001 ()2-s2.0-85099975118 (Scopus ID)
Available from: 2021-02-18 Created: 2021-02-18 Last updated: 2023-09-05Bibliographically approved
Igamberdiev, A. U. & Kleczkowski, L. A. (2021). Pyrophosphate as an alternative energy currency in plants. Biochemical Journal, 478(8), 1515-1524
Open this publication in new window or tab >>Pyrophosphate as an alternative energy currency in plants
2021 (English)In: Biochemical Journal, ISSN 0264-6021, E-ISSN 1470-8728, Vol. 478, no 8, p. 1515-1524Article, review/survey (Refereed) Published
Abstract [en]

In the conditions of [Mg2+] elevation that occur, in particular, under low oxygen stress and are the consequence of the decrease in [ATP] and increase in [ADP] and [AMP], pyrophosphate (PPi) can function as an alternative energy currency in plant cells. In addition to its production by various metabolic pathways, PPi can be synthesized in the combined reactions of pyruvate, phosphate dikinase (PPDK) and pyruvate kinase (PK) by so-called PK/PPDK substrate cycle, and in the reverse reaction of membrane-bound H+-pyrophosphatase, which uses the energy of electrochemical gradients generated on tonoplast and plasma membrane. The PPi can then be consumed in its active forms of MgPPi and Mg2PPi by PPi-utilizing enzymes, which require an elevated [Mg2+]. This ensures a continuous operation of glycolysis in the conditions of suppressed ATP synthesis, keeping metabolism energy efficient and less dependent on ATP.

Place, publisher, year, edition, pages
Portland Press, 2021
Keywords
adenylate equilibrium, adenylate kinase, free magnesium, membrane-bound h+-pyrophosphatase, pyrophosphate ppi, thermodynamic buffering
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-182928 (URN)10.1042/BCJ20200940 (DOI)000642604100003 ()2-s2.0-85104844307 (Scopus ID)
Available from: 2021-05-20 Created: 2021-05-20 Last updated: 2023-09-05Bibliographically approved
Kleczkowski, L. A. & Igamberdiev, A. U. (2020). Optimization of nucleotide sugar supply for polysaccharide formation via thermodynamic buffering. Biochemical Journal, 477(2), 341-356
Open this publication in new window or tab >>Optimization of nucleotide sugar supply for polysaccharide formation via thermodynamic buffering
2020 (English)In: Biochemical Journal, ISSN 0264-6021, E-ISSN 1470-8728, Vol. 477, no 2, p. 341-356Article in journal (Refereed) Published
Abstract [en]

Plant polysaccharides (cellulose, hemicellulose, pectin, starch) are either direct (i.e. leaf starch) or indirect products of photosynthesis, and they belong to the most abundant organic compounds in nature. Although each of these polymers is made by a specific enzymatic machinery, frequently in different cell locations, details of their synthesis share certain common features. Thus, the production of these polysaccharides is preceded by the formation of nucleotide sugars catalyzed by fully reversible reactions of various enzymes, mostly pyrophosphorylases. These 'buffering' enzymes are, generally, quite active and operate close to equilibrium. The nucleotide sugars are then used as substrates for irreversible reactions of various polysaccharide-synthesizing glycosyltransferases ('engine' enzymes), e.g. plastidial starch synthases, or plasma membrane-bound cellulose synthase and callose synthase, or ER/Golgi-located variety of glycosyltransferases forming hemicellulose and pectin backbones. Alternatively, the irreversible step might also be provided by a carrier transporting a given immediate precursor across a membrane. Here, we argue that local equilibria, established within metabolic pathways and cycles resulting in polysaccharide production, bring stability to the system via the arrangement of a flexible supply of nucleotide sugars. This metabolic system is itself under control of adenylate kinase and nucleoside-diphosphate kinase, which determine the availability of nucleotides (adenylates, uridylates, guanylates and cytidylates) and Mg2+, the latter serving as a feedback signal from the nucleotide metabolome. Under these conditions, the supply of nucleotide sugars to engine enzymes is stable and constant, and the metabolic process becomes optimized in its load and consumption, making the system steady and self-regulated.

Place, publisher, year, edition, pages
Portland Press, 2020
Keywords
adenylate kinase, ADP-glucose pyrophosphorylase, nucleoside-diphosphate kinase, starch synthesis, sucrose synthase, thermodynamic buffering
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-168571 (URN)10.1042/BCJ20190807 (DOI)000512948900003 ()31967651 (PubMedID)2-s2.0-85078309649 (Scopus ID)
Available from: 2020-03-05 Created: 2020-03-05 Last updated: 2023-03-24Bibliographically approved
Igamberdiev, A. U. & Kleczkowski, L. A. (2019). Thermodynamic buffering, stable non-equilibrium and establishment of the computable structure of plant metabolism. Progress in Biophysics and Molecular Biology, 146, 23-36
Open this publication in new window or tab >>Thermodynamic buffering, stable non-equilibrium and establishment of the computable structure of plant metabolism
2019 (English)In: Progress in Biophysics and Molecular Biology, ISSN 0079-6107, E-ISSN 1873-1732, Vol. 146, p. 23-36Article, review/survey (Refereed) Published
Abstract [en]

The equilibria of coenzyme nucleotides and substrates established in plant cells generate simple rules that govern the plant metabolome and provide optimal conditions for the non-equilibrium fluxes of major metabolic processes such as ATP synthesis, CO2 fixation, and mitochondrial respiration. Fast and abundant enzymes, such as adenylate kinase, carbonic anhydrase or malate dehydrogenase, provide constant substrate flux for these processes. These "buffering" enzymes follow the Michaelis-Menten (MM) kinetics and operate near equilibrium. The non-equilibrium "engine" enzymes, such as ATP synthase, Rubisco or the respiratory complexes, follow the modified version of MM kinetics due to their high concentration and low concentration of their substrates. The equilibrium reactions serve as control gates for the non-equilibrium flux through the engine enzymes establishing the balance of the fluxes of load and consumption of metabolic components. Under the coordinated operation of buffering and engine enzymes, the concentrations of free and Mg-bound adenylates and of free Mg2+ are set, serving as feedback signals from the adenylate metabolome. Those are linked to various cell energetics parameters, including membrane potentials. Also, internal levels of reduced and oxidized pyridine nucleotides are established in the coordinated operation of malate dehydrogenase and respiratory components, with proton concentration as a feedback from pyridine nucleotide pools. Non-coupled pathways of respiration serve to equilibrate the levels of pyridine nucleotides, adenylates, and as a pH stat. This stable non-equilibrium organizes the fluxes of energy spatially and temporally, controlling the rates of major metabolic fluxes that follow thermodynamically and kinetically defined computational principles. 

Place, publisher, year, edition, pages
PERGAMON-ELSEVIER SCIENCE LTD, 2019
Keywords
adenylates, Metabolomics, pH stat, Pyridine nucleotide, Stable non-equilibrium, Thermodynamic ffering, Uncoupling
National Category
Biochemistry and Molecular Biology Bioinformatics and Systems Biology
Identifiers
urn:nbn:se:umu:diva-164509 (URN)10.1016/j.pbiomolbio.2018.11.005 (DOI)000483909100003 ()30444975 (PubMedID)2-s2.0-85070651317 (Scopus ID)
Available from: 2019-11-22 Created: 2019-11-22 Last updated: 2023-03-24Bibliographically approved
Decker, D. & Kleczkowski, L. A. (2019). UDP-Sugar Producing Pyrophosphorylases: Distinct and Essential Enzymes With Overlapping Substrate Specificities, Providing de novo Precursors for Glycosylation Reactions. Frontiers in Plant Science, 9, Article ID 1822.
Open this publication in new window or tab >>UDP-Sugar Producing Pyrophosphorylases: Distinct and Essential Enzymes With Overlapping Substrate Specificities, Providing de novo Precursors for Glycosylation Reactions
2019 (English)In: Frontiers in Plant Science, E-ISSN 1664-462X, Vol. 9, article id 1822Article, review/survey (Refereed) Published
Abstract [en]

Nucleotide sugars are the key precursors for all glycosylation reactions and are required both for oligo- and polysaccharides synthesis and protein and lipid glycosylation. Among all nucleotide sugars, UDP-sugars are the most important precursors for biomass production in nature (e.g., synthesis of cellulose, hemicellulose, and pectins for cell wall production). Several recent studies have already suggested a potential role for UDP-Glc in plant growth and development, and UDP-Glc has also been suggested as a signaling molecule, in addition to its precursor function. In this review, we will cover primary mechanisms of formation of UDP-sugars, by focusing on UDP-sugar metabolizing pyrophosphorylases. The pyrophosphorylases can be divided into three families: UDP-Glc pyrophosphorylase (UGPase), UDP-sugar pyrophosphorylase (USPase), and UDP-N-acetyl glucosamine pyrophosphorylase (UAGPase), which can be distinguished both by their amino acid sequences and by differences in substrate specificity. Substrate specificities of these enzymes are discussed, along with structure-function relationships, based on their crystal structures and homology modeling. Earlier studies with transgenic plants have revealed that each of the pyrophosphorylases is essential for plant survival, and their loss or a decrease in activity results in reproductive impairment. This constitutes a problem when studying exact in vivo roles of the enzymes using classical reverse genetics approaches. Thus, strategies involving the use of specific inhibitors (reverse chemical genetics) are also discussed. Further characterization of the properties/roles of pyrophosphorylases should address fundamental questions dealing with mechanisms and control of carbohydrate synthesis and may allow to identify targets for manipulation of biomass production in plants.

Place, publisher, year, edition, pages
Frontiers Media S.A., 2019
Keywords
carbohydrate biosynthesis, chemical genetics, nucleotide sugar synthesis, enzyme substrate specificity, UDP-glucose pyrophosphorylase, UDP-N-acetylglucosamine pyrophosphorylase, UDP-sugar pyrophosphorylase
National Category
Botany
Identifiers
urn:nbn:se:umu:diva-155633 (URN)10.3389/fpls.2018.01822 (DOI)000454879300001 ()2-s2.0-85062809783 (Scopus ID)
Funder
Swedish Research CouncilThe Kempe FoundationsLars Hierta Memorial Foundation
Available from: 2019-01-25 Created: 2019-01-25 Last updated: 2024-01-17Bibliographically approved
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ORCID iD: ORCID iD iconorcid.org/0000-0001-8685-9665

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