scholarly journals Atomic structures of the RNA end-healing 5′-OH kinase and 2′,3′-cyclic phosphodiesterase domains of fungal tRNA ligase: conformational switches in the kinase upon binding of the GTP phosphate donor

2019 ◽  
Author(s):  
Ankan Banerjee ◽  
Yehuda Goldgur ◽  
Beate Schwer ◽  
Stewart Shuman
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Fungal Pathogens ◽  
Pathogenic Fungus ◽  
Atomic Structures ◽  
Polynucleotide Kinase ◽  
Phosphate Donor ◽  
Guanine Base ◽  
Conformational Switches ◽  
Β Sheet

Abstract Fungal tRNA ligase (Trl1) rectifies RNA breaks with 2′,3′-cyclic-PO4 and 5′-OH termini. Trl1 consists of three catalytic modules: an N-terminal ligase (LIG) domain; a central polynucleotide kinase (KIN) domain; and a C-terminal cyclic phosphodiesterase (CPD) domain. Trl1 enzymes found in all human fungal pathogens are untapped targets for antifungal drug discovery. Here we report a 1.9 Å crystal structure of Trl1 KIN-CPD from the pathogenic fungus Candida albicans, which adopts an extended conformation in which separate KIN and CPD domains are connected by an unstructured linker. CPD belongs to the 2H phosphotransferase superfamily by dint of its conserved central concave β sheet and interactions of its dual HxT motif histidines and threonines with phosphate in the active site. Additional active site motifs conserved among the fungal CPD clade of 2H enzymes are identified. We present structures of the Candida Trl1 KIN domain at 1.5 to 2.0 Å resolution—as apoenzyme and in complexes with GTP•Mg2+, IDP•PO4, and dGDP•PO4—that highlight conformational switches in the G-loop (which recognizes the guanine base) and lid-loop (poised over the nucleotide phosphates) that accompany nucleotide binding.

Control of Winged Waterprimrose (Jussiaea decurrens) and Northern Jointvetch (Aeschynomene virginica) with Fungal Pathogens

Weed Science ◽  
1979 ◽  
Vol 27 (5) ◽  
pp. 497-501 ◽  
Author(s):  
C. D. Boyette ◽  
G. E. Templeton ◽  
R. J. Smith
Keyword(s):  
Oryza Sativa ◽  
Glycine Max ◽  
Fungal Pathogens ◽  
Liquid Culture ◽  
Field Experiments ◽  
Colletotrichum Gloeosporioides ◽  
Pathogenic Fungus ◽  
Field Tests ◽  
Wide Range ◽  
Growing Region

An indigenous, host-specific, pathogenic fungus that parasitizes winged waterprimrose [Jussiaea decurrens(Walt.) DC.] is endemic in the rice growing region of Arkansas. The fungus was isolated and identified asColletotrichum gloeosporioides(Penz.) Sacc. f.sp. jussiaeae(CGJ). It is highly specific for parasitism of winged waterprimrose and not parasitic on creeping waterprimrose (J. repensL. var.glabrescensKtze.), rice (Oryza sativaL.), soybeans [Glycine max(L.) Merr.], cotton (Gossypium hirsutumL.), or 4 other crops and 13 other weeds. The fungus was physiologically distinct from C.gloeosporioides(Penz.) Sacc. f. sp.aeschynomene(CGA), an endemic anthracnose pathogen of northern jointvetch[Aeschynomene virginica(L.) B.S.P.], as indicated by cross inoculations of both weeds. Culture in the laboratory and inoculation of winged waterprimrose in greenhouse, growth chamber and field experiments indicated that the pathogen was stable, specific, and virulent in a wide range of environments. The pathogen yielded large quantities of spores in liquid culture. It is suitable for control of winged waterprimrose. Winged waterprimrose and northern jointvetch were controlled in greenhouse and field tests by application of spore mixtures of CGJ and CGA at concentrations of 1 to 2 million spores/ml of each fungus in 94 L/ha of water; the fungi did not damage rice or nontarget crops.

scholarly journals Crystal structures of non-oxidative decarboxylases reveal a new mechanism of action with a catalytic dyad and structural twists

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Matthias Zeug ◽  
Nebojsa Markovic ◽  
Cristina V. Iancu ◽  
Joanna Tripp ◽  
Mislav Oreb ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Active Site ◽  
Enzyme Engineering ◽  
Base Catalysis ◽  
Oxidative Decarboxylation ◽  
Transition State Stabilization ◽  
Major Bottleneck ◽  
Hydroxybenzoic Acids ◽  
Potassium Binding ◽  
Β Sheet

AbstractHydroxybenzoic acids, like gallic acid and protocatechuic acid, are highly abundant natural compounds. In biotechnology, they serve as critical precursors for various molecules in heterologous production pathways, but a major bottleneck is these acids’ non-oxidative decarboxylation to hydroxybenzenes. Optimizing this step by pathway and enzyme engineering is tedious, partly because of the complicating cofactor dependencies of the commonly used prFMN-dependent decarboxylases. Here, we report the crystal structures (1.5–1.9 Å) of two homologous fungal decarboxylases, AGDC1 from Arxula adenivorans, and PPP2 from Madurella mycetomatis. Remarkably, both decarboxylases are cofactor independent and are superior to prFMN-dependent decarboxylases when heterologously expressed in Saccharomyces cerevisiae. The organization of their active site, together with mutational studies, suggests a novel decarboxylation mechanism that combines acid–base catalysis and transition state stabilization. Both enzymes are trimers, with a central potassium binding site. In each monomer, potassium introduces a local twist in a β-sheet close to the active site, which primes the critical H86-D40 dyad for catalysis. A conserved pair of tryptophans, W35 and W61, acts like a clamp that destabilizes the substrate by twisting its carboxyl group relative to the phenol moiety. These findings reveal AGDC1 and PPP2 as founding members of a so far overlooked group of cofactor independent decarboxylases and suggest strategies to engineer their unique chemistry for a wide variety of biotechnological applications.

scholarly journals Crystal Structure, Exogenous Ligand Binding, and Redox Properties of an Engineered Diiron Active Site in a Bacterial Hemerythrin

2013 ◽  
Vol 52 (22) ◽  
pp. 13014-13020 ◽  
Author(s):  
Yasunori Okamoto ◽  
Akira Onoda ◽  
Hiroshi Sugimoto ◽  
Yu Takano ◽  
Shun Hirota ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Ligand Binding ◽  
Active Site ◽  
Redox Properties ◽  
Exogenous Ligand

scholarly journals Crystal structure of Pistol, a class of self-cleaving ribozyme

2017 ◽  
Vol 114 (5) ◽  
pp. 1021-1026 ◽  
Author(s):  
Laura A. Nguyen ◽  
Jimin Wang ◽  
Thomas A. Steitz
Keyword(s):  
Crystal Structure ◽  
Rna Structure ◽  
Tertiary Structure ◽  
Genomic Analysis ◽  
Comparative Genomic ◽  
Nonbridging Oxygen ◽  
Close Proximity ◽  
Domains Of Life ◽  
Guanine Base ◽  
A Minor

Small self-cleaving ribozymes have been discovered in all evolutionary domains of life. They can catalyze site-specific RNA cleavage, and as a result, they have relevance in gene regulation. Comparative genomic analysis has led to the discovery of a new class of small self-cleaving ribozymes named Pistol. We report the crystal structure of Pistol at 2.97-Å resolution. Our results suggest that the Pistol ribozyme self-cleavage mechanism likely uses a guanine base in the active site pocket to carry out the phosphoester transfer reaction. The guanine G40 is in close proximity to serve as the general base for activating the nucleophile by deprotonating the 2′-hydroxyl to initiate the reaction (phosphoester transfer). Furthermore, G40 can also establish hydrogen bonding interactions with the nonbridging oxygen of the scissile phosphate. The proximity of G32 to the O5′ leaving group suggests that G32 may putatively serve as the general acid. The RNA structure of Pistol also contains A-minor interactions, which seem to be important to maintain its tertiary structure and compact fold. Our findings expand the repertoire of ribozyme structures and highlight the conserved evolutionary mechanism used by ribozymes for catalysis.

scholarly journals Crystal Structure of the Dithiol Oxidase DsbA Enzyme fromProteus MirabilisBound Non-covalently to an Active Site Peptide Ligand

2014 ◽  
Vol 289 (28) ◽  
pp. 19810-19822 ◽  
Author(s):  
Fabian Kurth ◽  
Wilko Duprez ◽  
Lakshmanane Premkumar ◽  
Mark A. Schembri ◽  
David P. Fairlie ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Active Site ◽  
Peptide Ligand

WHICH IS THE PROTON-SHUTTLE IN ISOXANTHOPTERIN DEAMINASE? QM/MM MD UNDERSTANDING

2013 ◽  
Vol 12 (08) ◽  
pp. 1341002 ◽  
Author(s):  
XIN ZHANG ◽  
MING LEI
Keyword(s):  
Crystal Structure ◽  
Molecular Dynamics ◽  
Hydrogen Bonds ◽  
Proton Transfer ◽  
Glutamic Acid ◽  
Active Site ◽  
Nucleophilic Attack ◽  
Zinc Ions ◽  
Initial Model ◽  
Dynamics Simulations

The deamination process of isoxanthopterin catalyzed by isoxanthopterin deaminase was determined using the combined QM(PM3)/MM molecular dynamics simulations. In this paper, the updated PM3 parameters were employed for zinc ions and the initial model was built up based on the crystal structure. Proton transfer and following steps have been investigated in two paths: Asp336 and His285 serve as the proton shuttle, respectively. Our simulations showed that His285 is more effective than Aap336 in proton transfer for deamination of isoxanthopterin. As hydrogen bonds between the substrate and surrounding residues play a key role in nucleophilic attack, we suggested mutating Thr195 to glutamic acid, which could enhance the hydrogen bonds and help isoxanthopterin get close to the active site. The simulations which change the substrate to pterin 6-carboxylate also performed for comparison. Our results provide reference for understanding of the mechanism of deaminase and for enhancing the deamination rate of isoxanthopterin deaminase.

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