Bacterial Glycogen Synthesis and Its Regulation

Preiss, Jack

doi:10.1146/annurev.mi.38.100184.002223

Cited by 369 publications

(143 citation statements)

References 58 publications

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“…To expedite this process, we developed a program in Pascal language. 2 This modified method of hydrophobic cluster analysis proved to be more accurate in finding similar clusters in these homologous proteins.…”

Section: Structure Prediction Analysismentioning

confidence: 96%

Aspartate Residue 142 Is Important for Catalysis by ADP-glucose Pyrophosphorylase from Escherichia coli

Frueauf

Ballícora

Preiss

2001

Journal of Biological Chemistry

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Glycogen and starch, which consist of mainly ␣1,4-polyglucans, encompass most of the energy reserves in organisms. The biosynthesis of these polysaccharides proceeds from the formation of the glucosyl donor, ADP-glucose (ADP-Glc).1 ADP-glucose pyrophosphorylase (ADP-Glc PPase, EC 2.7.7.27) catalyzes the synthesis of ADP-Glc and pyrophosphate (PP i ) from ATP and glucose 1-phosphate (Glc-1-P) (1). This reversible reaction is the key regulatory step in the production of glycogen in bacteria and starch in plants (2-5).ADP-Glc PPase has been isolated and characterized from various sources. Most of the enzymes are allosterically regulated by the glycolytic intermediates of the major pathway of carbon assimilation in the organism (6). The regulatory effectors include the activator fructose 1,6-bisphosphate (FBP) and the inhibitor AMP for an enzyme from enteric bacteria, e.g. Escherichia coli (7), and the activator 3-phosphoglycerate and the inhibitor inorganic phosphate for enzymes from plants and other photosynthetic organisms, e.g. algae and cyanobacteria (4, 5). Active ADP-Glc PPase forms a tetramer of about 200 kDa, which is either homomeric (␣ 4 ) in bacteria (8, 9) or heteromeric (␣ 2 ␤ 2 ) in plants (10, 11). The emergence of two different subunits in plants, designated as small and large based on disparities in molecular weight, likely arose from the need for tissue-specific regulation (5, 12). Also, other studies show that each subunit has a separate function: the small subunit is catalytic while the large subunit is a mediator of the allosteric regulation of the small subunit (13).The three-dimensional structure of an ADP-Glc PPase has not yet been resolved by x-ray crystallography; however, many structural studies have been performed using other approaches. A combination of techniques, such as chemical modification and site-directed mutagenesis, has been applied to study the ADP-Glc PPase enzyme from E. coli. Azido-based photoaffinity analogs of the ATP substrate and the ADP-Glc substrate (pyrophosphorolysis direction) were used to probe the substrate-binding site. Analysis of the covalently labeled enzyme revealed Tyr 114 as the modified residue, and subsequent site-directed mutagenesis of the residue showed a marked decrease in affinity for ATP, as well as a decrease in affinity for . The residue likely interacts with the adenine ring of ATP but is not crucial for the binding of the substrate. Pyridoxal-5-phosphate, a mild reagent that modifies lysines within ligand-binding sites upon reduction by NaBH 4 , is an analog of the activator from E. coli, FBP. Reaction of this modifying agent with the enzyme labels two different lysines, Lys 39 and Lys 195 that are blocked by the inclusion of FBP and of the substrate ADP-Glc, respectively (15, 16). Mutation of Lys 39 showed that this residue is important in the interaction of the activator FBP with the enzyme (17). Mutagenesis of the Lys 195 residue produced enzymes with drastically increased K m values for Glc-1-P, while the other kinetic constants and the k c...

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Section: Structure Prediction Analysismentioning

confidence: 96%

Aspartate Residue 142 Is Important for Catalysis by ADP-glucose Pyrophosphorylase from Escherichia coli

Frueauf

Ballícora

Preiss

2001

Journal of Biological Chemistry

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“…Besides, ApS2 mRNA can only be detected by reverse transcription-PCR, indicating that its level of expression is very low. 2 So, it is possible that APS2 is a noncatalytic subunit. Nevertheless, further studies are required to confirm this hypothesis and discard other possible roles of the protein.…”

Section: Fig 2 Activation By 3-pga Of the Recombinant Arabidopsis Amentioning

confidence: 99%

“…way (1)(2)(3). It catalyzes the synthesis of ADP-glucose (ADP-Glc) and PP i from glucose 1-phosphate (Glc-1-P) and ATP (4).…”

mentioning

confidence: 99%

The Different Large Subunit Isoforms of Arabidopsis thaliana ADP-glucose Pyrophosphorylase Confer Distinct Kinetic and Regulatory Properties to the Heterotetrameric Enzyme

Crevillén

Ballícora

Mérida

et al. 2003

Journal of Biological Chemistry

126

153

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ADP-glucose pyrophosphorylase catalyzes the first and limiting step in starch biosynthesis and is allosterically regulated by the levels of 3-phosphoglycerate and phosphate in plants. ADP-glucose pyrophosphorylases from plants are heterotetramers composed of two types of subunits (small and large). In this study, the six Arabidopsis thaliana genes coding for ADP-glucose pyrophosphorylase isoforms (two small and four large subunits) have been cloned and expressed in an Escherichia coli mutant deficient in ADP-glucose pyrophosphorylase activity. The co-expression of the small subunit APS1 with the different Arabidopsis large subunits (APL1, APL2, APL3, and APL4) resulted in heterotetramers with different regulatory and kinetic properties. Heterotetramers composed of APS1 and APL1 showed the highest sensitivity to the allosteric effectors as well as the highest apparent affinity for the substrates (glucose-1-phosphate and ATP), whereas heterotetramers formed by APS1 and APL2 showed the lower response to allosteric effectors and the lower affinity for the substrates. No activity was detected for the second gene coding for a small subunit isoform (APS2) annotated in the Arabidopsis genome. This lack of activity is possibly due to the absence of essential amino acids involved in catalysis and/or in the binding of glucose-1-phosphate and 3-phosphoglycerate. Kinetic and regulatory properties of the different heterotetramers, together with sequence analysis has allowed us to make a distinction between sink and source enzymes, because the combination of different large subunits would provide a high plasticity to ADP-glucose pyrophosphorylase activity and regulation. This is the first experimental data concerning the role that all the ADP-glucose pyrophosphorylase isoforms play in a single plant species. This phenomenon could have an important role in vivo, because different large subunits would confer distinct regulatory properties to ADP-glucose pyrophosphorylase according to the necessities for starch synthesis in a given tissue.

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“…The fold activation of Devo 202, Devo 330, and Devo 339 is low for all metabolites because of an increased level of activity in the absence of activator, whereas Devo 206 and Devo 316 exhibit meaningful fold activation by 3-PGA and Fru-1,6-bisP, respectively, while having significant activity in the absence of activator. The AGPases from bacteria, blue-green algae, and higher plants can be divided into seven groups based on differences in activator specificity (1). Six groups prefer pyruvate, Fru-6-P, Fru-1,6-bisP, 3-PGA, pyridoxal phosphate, or NADPH as an effector, with one group showing no activation by any known metabolite.…”

Section: Sensitivity and Selectivity Of Devo Mutants To Metabolite Acmentioning

confidence: 99%

“…The catalytic activity of the enzyme is modulated by small effector molecules whose nature reflects the major carbon assimilatory pathway of the organism. Many bacterial enzymes are activated by intermediates of glycolysis or the Entner Douderoff pathway [e.g., pyruvate, fructose-6-phosphate (Fru-6-P), fructose-1,6-bisphosphate (Fru-1,6-bisP 2 )] and inhibited by AMP (1). The enzymes from blue-green algae and higher plants are activated by 3-phosphoglyceric acid and inhibited by Pi, key intermediates in CO 2 assimilation by the C 3 pathway.…”

mentioning

confidence: 99%

Directed molecular evolution of ADP-glucose pyrophosphorylase

Salamone

Kavaklı

Slattery

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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ADP-glucose pyrophosphorylase catalyzes a rate-limiting reaction in prokaryotic glycogen and plant starch biosynthesis. Despite sharing similar molecular size and catalytic and allosteric regulatory properties, the prokaryotic and higher plant enzymes differ in higher-order protein structure. The bacterial enzyme is encoded by a single gene whose product of ca. 50,000 Da assembles into a homotetrameric structure. Although the higher plant enzyme has a similar molecular size, it is made up of a pair of large subunits and a pair of small subunits, encoded by different genes. To identify the basis for the evolution of AGPase function and quaternary structure, a potato small subunit homotetrameric mutant, TG-15, was subjected to iterations of DNA shuffling and screened for enzyme variants with up-regulated catalytic and͞or regulatory properties. A glycogen selection͞screening regimen of buoyant density gradient centrifugation and iodine vapor colony staining on glucosecontaining media was used to increase the stringency of selection. This approach led to the isolation of a population of AGPase small subunit homotetramer enzymes with enhanced affinity toward ATP and increased sensitivity to activator and͞or greater resistance to inhibition than TG-15. Several enzymes displayed a shift in effector preference from 3-phosphoglycerate to fructose-6 phosphate or fructose-1,6-bis-phosphate, effectors used by specific bacterial AGPases. Our results suggest that evolution of AGPase, with regard to quaternary structure, allosteric effector selectivity, and effector sensitivity, can occur through the introduction of a few point mutations alone with low-level recombination hastening the process.starch ͉ metabolic engineering ͉ allosteric regulation ͉ DNA shuffling

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Bacterial Glycogen Synthesis and Its Regulation

Cited by 369 publications

References 58 publications

Aspartate Residue 142 Is Important for Catalysis by ADP-glucose Pyrophosphorylase from Escherichia coli

Aspartate Residue 142 Is Important for Catalysis by ADP-glucose Pyrophosphorylase from Escherichia coli

The Different Large Subunit Isoforms of Arabidopsis thaliana ADP-glucose Pyrophosphorylase Confer Distinct Kinetic and Regulatory Properties to the Heterotetrameric Enzyme

Directed molecular evolution of ADP-glucose pyrophosphorylase

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