The Complex of
            <i>Sphingomonas elodea</i>
            ATCC 31461 Glucose-1-Phosphate Uridylyltransferase with Glucose-1-Phosphate Reveals a Novel Quaternary Structure, Unique among Nucleoside Diphosphate-Sugar Pyrophosphorylase Members

Aragão, D.; Fialho, Arsénio M.; Marques, Ana Rita; Mitchell, Edward P.; Sá‐Correia, Isabel; Frazão, Carlos

doi:10.1128/jb.00277-07

Cited by 31 publications

(32 citation statements)

References 33 publications

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“…Since this paper was accepted, another manuscript appeared online describing the structure of UGPase from Sphingomonas elodea complexed with glucose-1-phosphate (Aragão et al 2007). This structure, in combination with our results, provides a more detailed understanding of the active site geometry for the UGPases.…”

Section: Note Added In Proofmentioning

confidence: 99%

Active site geometry of glucose‐1‐phosphate uridylyltransferase

Thoden

Holden

2007

Protein Science

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Glucose-1-phosphate uridylyltransferase, or UGPase, catalyzes the production of UDP-glucose from glucose-1-phosphate and UTP. Because of the biological role of UDP-glucose in glycogen synthesis and in the formation of glycolipids, glycoproteins, and proteoglycans, the enzyme is widespread in nature. Recently this laboratory reported the three-dimensional structure of UGPase from Escherichia coli. While the initial X-ray analysis revealed the overall fold of the enzyme, details concerning its active site geometry were limited because crystals of the protein complexed with either substrates or products could never be obtained. In an effort to more fully investigate the active site geometry of the enzyme, UGPase from Corynebacterium glutamicum was subsequently cloned and purified. Here we report the X-ray structure of UGPase crystallized in the presence of both magnesium and UDP-glucose. Residues involved in anchoring the ligand to the active site include the polypeptide chain backbone atoms of Ala 20, Gly 21, Gly 117, Gly 180, and Ala 214, and the side chains of Glu 36, Gln 112, Asp 143, Glu 201, and Lys 202. Two magnesium ions are observed coordinated to the UDP-glucose. An a-and a bphosphoryl oxygen, three waters, and the side chain of Asp 142 ligate the first magnesium, whereas the second ion is coordinated by an a-phosphoryl oxygen and five waters. The position of the first magnesium is conserved in both the glucose-1-phosphate thymidylyltransferases and the cytidylyltransferases. The structure presented here provides further support for the role of the conserved magnesium ion in the catalytic mechanisms of the sugar-1-phosphate nucleotidylyltransferases.

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Section: Note Added In Proofmentioning

confidence: 99%

Active site geometry of glucose‐1‐phosphate uridylyltransferase

Thoden

Holden

2007

Protein Science

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“…6,17 The structures of UGPs from several organisms have been elucidated and reveal a common structural arrangement. [26][27][28][29][30][31] These eukaryotic UGPs adopt a tripartite structure with a small N-terminal domain, a central catalytic domain having a Rossmann-like α/β/α sandwich fold, and a β-helix C-terminal domain. [27][28][29] In fungi and mammals, the latter domain is involved in oligomerization, whereas L. major and plant UGPs are active as monomers.…”

Section: Introductionmentioning

confidence: 99%

Structural Basis for the Broad Substrate Range of the UDP-Sugar Pyrophosphorylase from Leishmania major

Dickmanns

Damerow

Neumann

et al. 2011

Journal of Molecular Biology

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“…Eukaryotic UDP-Glc PPases are bigger than those found in bacteria [5,6], and the enzyme from Entamoeba histolytica (and probably from all protozoa) was recently characterized as being regulated by redox modification of critical cysteinyl residues [7]. Many bacterial UDP-Glc PPases have been characterized [8–13], and the crystallographic structures of the enzyme from Escherichia coli [14], Sphingomonas elodea [15] and Corynebacterium glutamicum [16] have been elucidated. The prokaryotic UDP-Glc PPase is a dimeric/tetrameric protein formed by a single subunit of ~35 kDa with a relatively high specific activity and specificity for Glc-1P and UTP [8–13].…”

Section: Introductionmentioning

confidence: 99%

A Chimeric UDP-Glucose Pyrophosphorylase Produced by Protein Engineering Exhibits Sensitivity to Allosteric Regulators

Diez

Ebrecht

Martínez

et al. 2013

IJMS

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In bacteria, glycogen or oligosaccharide accumulation involves glucose-1-phosphate partitioning into either ADP-glucose (ADP-Glc) or UDP-Glc. Their respective synthesis is catalyzed by allosterically regulated ADP-Glc pyrophosphorylase (EC 2.7.7.27, ADP-Glc PPase) or unregulated UDP-Glc PPase (EC 2.7.7.9). In this work, we characterized the UDP-Glc PPase from Streptococcus mutans. In addition, we constructed a chimeric protein by cutting the C-terminal domain of the ADP-Glc PPase from Escherichia coli and pasting it to the entire S. mutans UDP-Glc PPase. Both proteins were fully active as UDP-Glc PPases and their kinetic parameters were measured. The chimeric enzyme had a slightly higher affinity for substrates than the native S. mutans UDP-Glc PPase, but the maximal activity was four times lower. Interestingly, the chimeric protein was sensitive to regulation by pyruvate, 3-phosphoglyceric acid and fructose-1,6-bis-phosphate, which are known to be effectors of ADP-Glc PPases from different sources. The three compounds activated the chimeric enzyme up to three-fold, and increased the affinity for substrates. This chimeric protein is the first reported UDP-Glc PPase with allosteric regulatory properties. In addition, this is a pioneer work dealing with a chimeric enzyme constructed as a hybrid of two pyrophosphorylases with different specificity toward nucleoside-diphospho-glucose and our results turn to be relevant for a deeper understanding of the evolution of allosterism in this family of enzymes.

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The Complex of Sphingomonas elodea ATCC 31461 Glucose-1-Phosphate Uridylyltransferase with Glucose-1-Phosphate Reveals a Novel Quaternary Structure, Unique among Nucleoside Diphosphate-Sugar Pyrophosphorylase Members

Cited by 31 publications

References 33 publications

Active site geometry of glucose‐1‐phosphate uridylyltransferase

Active site geometry of glucose‐1‐phosphate uridylyltransferase

Structural Basis for the Broad Substrate Range of the UDP-Sugar Pyrophosphorylase from Leishmania major

A Chimeric UDP-Glucose Pyrophosphorylase Produced by Protein Engineering Exhibits Sensitivity to Allosteric Regulators

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