New Determinants in the Catalytic Mechanism of Nucleoside Hydrolases from the Structures of Two Isozymes from Sulfolobus solfataricus

Biochemistry ◽  
2012 ◽  
Vol 51 (22) ◽  
pp. 4590-4599 ◽  
Author(s):  
Claudia Minici ◽  
Giovanna Cacciapuoti ◽  
Ester De Leo ◽  
Marina Porcelli ◽  
Massimo Degano
Keyword(s):  
Catalytic Mechanism ◽  
Sulfolobus Solfataricus ◽  
Nucleoside Hydrolases

Exploring the catalytic mechanism of the first dimeric Bcp: Functional, structural and docking analyses of Bcp4 from Sulfolobus solfataricus

Biochimie ◽  
2010 ◽  
Vol 92 (10) ◽  
pp. 1435-1444 ◽  
Author(s):  
Danila Limauro ◽  
Katia D’Ambrosio ◽  
Emma Langella ◽  
Giuseppina De Simone ◽  
Ilaria Galdi ◽  
...  
Keyword(s):  
Catalytic Mechanism ◽  
Sulfolobus Solfataricus

Thermal Unfolding of Nucleoside Hydrolases from the Hyperthermophilic Archaeon Sulfolobus solfataricus: Role of Disulfide Bonds

2012 ◽  
Vol 19 (3) ◽  
pp. 369-374 ◽  
Author(s):  
Marina Porcelli ◽  
Ester De Leo ◽  
Pompea Del Vecchio ◽  
Francesca Fuccio ◽  
Giovanna Cacciapuoti
Keyword(s):  
Disulfide Bonds ◽  
Sulfolobus Solfataricus ◽  
Thermal Unfolding ◽  
Hyperthermophilic Archaeon ◽  
Nucleoside Hydrolases

scholarly journals An extremely thermostable aldolase from Sulfolobus solfataricus with specificity for non-phosphorylated substrates

1999 ◽  
Vol 343 (3) ◽  
pp. 563-570 ◽  
Author(s):  
Catriona L. BUCHANAN ◽  
Helen CONNARIS ◽  
Michael J. DANSON ◽  
Christopher D. REEVE ◽  
David W. HOUGH
Keyword(s):  
Catalytic Mechanism ◽  
Sequence Similarity ◽  
Sulfolobus Solfataricus ◽  
Kinetic Properties ◽  
Cloning And Sequencing ◽  
Gene Encoding ◽  
Expression In Escherichia Coli ◽  
Carbon Carbon Bond ◽  
Step Procedure ◽  
Dependent Inactivation

Sulfolobus solfataricus is a hyperthermophilic archaeon growing optimally at 80-85 °C. It metabolizes glucose via a novel non-phosphorylated Entner-Doudoroff pathway, in which the reversible C6 to C3 aldol cleavage is catalysed by 2-keto-3-deoxygluconate aldolase (KDG-aldolase), generating pyruvate and glyceraldehyde. Given the ability of such a hyperstable enzyme to catalyse carbon-carbon-bond synthesis with non-phosphorylated metabolites, we report here the cloning and sequencing of the S. solfataricus gene encoding KDG-aldolase, and its expression in Escherichia coli to give fully active enzyme. The recombinant enzyme was purified in a simple two-step procedure, and shown to possess kinetic properties indistinguishable from the enzyme purified from S. solfataricuscells. The KDG-aldolase is a thermostable tetrameric protein with a half-life at 100 °C of 2.5 h, and is equally active with both D- and L-glyceraldehyde. It exhibits sequence similarity to the N-acetylneuraminate lyase superfamily of Schiff-base-dependent aldolases, dehydratases and decarboxylases, and evidence is presented for a similar catalytic mechanism for the archaeal enzyme by substrate-dependent inactivation by reduction with NaBH4.

Structures of c-di-GMP/cGAMP degrading phosphodiesterase VcEAL: identification of a novel conformational switch and its implication

2019 ◽  
Vol 476 (21) ◽  
pp. 3333-3353 ◽  
Author(s):  
Malti Yadav ◽  
Kamalendu Pal ◽  
Udayaditya Sen
Keyword(s):  
Active Site ◽  
Metal Binding ◽  
Metal Ion ◽  
Triosephosphate Isomerase ◽  
Catalytic Mechanism ◽  
Degradation Mechanism ◽  
Structural Basis ◽  
Conserved Residues ◽  
Phosphodiester Bond ◽  
Cyclic Dinucleotides

Cyclic dinucleotides (CDNs) have emerged as the central molecules that aid bacteria to adapt and thrive in changing environmental conditions. Therefore, tight regulation of intracellular CDN concentration by counteracting the action of dinucleotide cyclases and phosphodiesterases (PDEs) is critical. Here, we demonstrate that a putative stand-alone EAL domain PDE from Vibrio cholerae (VcEAL) is capable to degrade both the second messenger c-di-GMP and hybrid 3′3′-cyclic GMP–AMP (cGAMP). To unveil their degradation mechanism, we have determined high-resolution crystal structures of VcEAL with Ca2+, c-di-GMP-Ca2+, 5′-pGpG-Ca2+ and cGAMP-Ca2+, the latter provides the first structural basis of cGAMP hydrolysis. Structural studies reveal a typical triosephosphate isomerase barrel-fold with substrate c-di-GMP/cGAMP bound in an extended conformation. Highly conserved residues specifically bind the guanine base of c-di-GMP/cGAMP in the G2 site while the semi-conserved nature of residues at the G1 site could act as a specificity determinant. Two metal ions, co-ordinated with six stubbornly conserved residues and two non-bridging scissile phosphate oxygens of c-di-GMP/cGAMP, activate a water molecule for an in-line attack on the phosphodiester bond, supporting two-metal ion-based catalytic mechanism. PDE activity and biofilm assays of several prudently designed mutants collectively demonstrate that VcEAL active site is charge and size optimized. Intriguingly, in VcEAL-5′-pGpG-Ca2+ structure, β5–α5 loop adopts a novel conformation that along with conserved E131 creates a new metal-binding site. This novel conformation along with several subtle changes in the active site designate VcEAL-5′-pGpG-Ca2+ structure quite different from other 5′-pGpG bound structures reported earlier.

Exemplar Abstract for Sulfolobus solfataricus Zillig et al. 1980.

2003 ◽  
Author(s):  
Charles Thomas Parker ◽  
Dorothea Taylor ◽  
George M Garrity
Keyword(s):  
Sulfolobus Solfataricus

Rationalizing the pH-Activity Response for Escherichia Coli’s Glycinamide Ribonucleotidetransformylase Through Computational Methods

2019 ◽  
Author(s):  
Adrian Roitberg ◽  
Pancham Lal Gupta
Keyword(s):  
Transfer Reaction ◽  
Catalytic Mechanism ◽  
Ph Dependence ◽  
Ph Effects ◽  
Dependent Manner ◽  
E Coli ◽  
Gar Tfase ◽  
Activity Curve ◽  
Ph Dependent ◽  
Formyl Transfer

<div>Human Glycinamide ribonucleotide transformylase (GAR Tfase), a regulatory enzyme in the de novo purine biosynthesis pathway, has been established as an anti-cancer target. GAR Tfase catalyzes the formyl transfer reaction from the folate cofactor to the GAR ligand. In the present work, we study E. coli GAR Tfase, which has high sequence similarity with the human GAR Tfase with most functional residues conserved. E. coli GAR Tfase exhibits structural changes and the binding of ligands that varies with pH which leads to change the rate of the formyl transfer reaction in a pH-dependent manner. Thus, the inclusion of pH becomes essential for the study of its catalytic mechanism. Experimentally, the pH-dependence of the kinetic parameter kcat is measured to evaluate the pH-range of enzymatic activity. However, insufficient information about residues governing the pH-effects on the catalytic activity leads to ambiguous assignments of the general acid and base catalysts and consequently its catalytic mechanism. In the present work, we use pH-replica exchange molecular dynamics (pH-REMD) simulations to study the effects of pH on E. coli GAR Tfase enzyme. We identify the titratable residues governing the pH-dependent conformational changes in the system. Furthermore, we filter out the protonation states which are essential in maintaining the structural integrity, keeping the ligands bound and assisting the catalysis. We reproduce the experimental pH-activity curve by computing the population of key protonation states. Moreover, we provide a detailed description of residues governing the acidic and basic limbs of the pH-activity curve.</div>

Arylboronic Acids in Self-condensation of Glucosamines Forming Deoxyfructosazine, Catalysts or Reagents?

2019 ◽  
Author(s):  
Meifeng Wang ◽  
Gan Zhu ◽  
Yiqun Li ◽  
Liuqun Gu
Keyword(s):  
Molecular Recognition ◽  
Boric Acid ◽  
Boronic Acid ◽  
Catalytic Mechanism ◽  
Organic Transformations ◽  
Food Industries ◽  
Arylboronic Acids ◽  
Excess Amount ◽  
Phenyl Boronic Acid ◽  
Boron Acid

Arylboronic acids were widely used as efficient catalysts in direct amide formation and other organic transformations. Surprisingly, reports on their use as catalysts in carbohydrates synthesis are very rare even though boron acid-diol complexation was extensively investigated in molecular recognition for saccharides and so on. Here we developed an efficient arylboronic acids catalyzed dimerization of glucosamines forming deoxyfructosazine which is important compound in pharmaceutical and food industries, against a commonly held belief that excess amount of phenyl boronic acid (or boric acid) is a must. A catalytic mechanism was also proposed and arylboronic acids instead of their boronates was identified as catalysts.

Bifluoride Ion Mediated SuFEx Trifluoromethylation of Sulfonyl Fluorides and Iminosulfur Oxydifluorides

2018 ◽  
Author(s):  
Christopher J. Smedley ◽  
Bing Gao ◽  
Suhua Li ◽  
Qinheng Zheng ◽  
Andrew Molino ◽  
...  
Keyword(s):  
Breast Cancer Cells ◽  
Catalytic Mechanism ◽  
Bioactive Molecules ◽  
Chromatographic Purification ◽  
Rapid Exchange ◽  
Nucleophilic Displacement ◽  
Sulfur Fluoride ◽  
Reactions With Nucleophiles ◽  
New Generation ◽  
Selection Of

Sulfur-Fluoride Exchange (SuFEx) is the new generation click chemistry transformation exploiting the unique properties of S-F bonds and their ability to undergo near-perfect reactions with nucleophiles. We report here the first SuFEx based protocol for the efficient synthesis of pharmaceutically important triflones and bis(trifluoromethyl)sulfur oxyimines from the corresponding sulfonyl fluorides and iminosulfur oxydifluorides, respectively. The new protocol involves the rapid exchange of the S-F bond with trifluoromethyltrimethylsilane (TMSCF<sub>3</sub>) upon activation with potassium bifluoride in anhydrous DMSO. The reaction tolerates a wide selection of substrates and proceeds under mild conditions without need for chromatographic purification. A tentative catalytic mechanism is proposed supported by DFT calculations, involving formation of the free trifluoromethyl anion followed by nucleophilic displacement of the S-F through a five-coordinate intermediate. The preparation of a benzothiazole derived bis(trifluoromethyl)sulfur oxyimine with cytotoxic selectivity for MCF7 breast cancer cells demonstrates the utility of this methodology for the late-stage functionalization of bioactive molecules.<br>

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