scholarly journals An unusual metal ion configuration in a viral DNA-packaging nuclease active site

2014 ◽  
Vol 70 (a1) ◽  
pp. C489-C489
Author(s):  
Haiyan Zhao ◽  
Theodore Christensen ◽  
Zihan Lin ◽  
Annie Lynn ◽  
Liang Tang
Keyword(s):  
Nucleic Acid ◽  
Metal Ions ◽  
Active Site ◽  
Short Distance ◽  
Metal Ion ◽  
Catalytic Mechanism ◽  
Dna Packaging ◽  
Viral Dna ◽  
Viral Dna Packaging ◽  
Ion Cluster

Nucleic acid metabolism is fundamental to many biological processes. A large class of enzymes such as RNase H, reverse transcriptase, retroviral integrase, topoisomerase, DNA and RNA polymerase, transposase, Holliday-junction resolvase, RNAi slicer Argonaute, and viral DNA-packaging terminase, utilize a common two-metal-ion catalytic mechanism for cleavage or synthesis of nucleic acid chains. Here we report an unusual metal-ion cluster in the active site of the nuclease domain of a viral DNA-packaging terminase unveiled by X-ray structures up to 1.38 Angstrom resolution. Two Mg2+ ions are situated in a coupled octahedral coordination system with liganding oxygen atoms from aspartic acid residues as well as water molecules. The two Mg2+ ions are located within a strikingly short distance of ~2.5 Å, which is unusual given the 1.6 Å atomic radius of Mg2+ and is shorter than previously observed metal-metal distances in metallocluster-containing enzymes or other biological systems. This provides the structural basis for distinguishing Mg2+ from other metal ions such as Ca2+ which are well known to support binding of the nucleic acid substrate but not support catalysis. Such an ultra-short distance between two metal-ions may be essential for generation of a highly positive niche, leading to nucleophilic attack at the phosphodiester bond of DNA. These results have defined the precise chemical configuration of the active site in nucleases using two-metal-ion catalytic mechanism. Moreover, assembly of this two-metal-ion cluster in the viral DNA-packaging terminase is mediated by an adjacent Lys residue, likely serving as a regulatory mechanism for activation of the nuclease activity of the terminase during packaging of viral genome.

scholarly journals Two distinct modes of metal ion binding in the nuclease active site of a viral DNA-packaging terminase: insight into the two-metal-ion catalytic mechanism

2015 ◽  
Vol 43 (22) ◽  
pp. 11003-11016 ◽  
Author(s):  
Haiyan Zhao ◽  
Zihan Lin ◽  
Anna Y. Lynn ◽  
Brittany Varnado ◽  
John A. Beutler ◽  
...  
Keyword(s):  
Active Site ◽  
Metal Ion ◽  
Catalytic Mechanism ◽  
Dna Packaging ◽  
Ion Binding ◽  
Metal Ion Binding ◽  
Viral Dna ◽  
Viral Dna Packaging ◽  
Insight Into

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.

Na+/K+exchange switches the catalytic apparatus of potassium-dependent plantL-asparaginase

2014 ◽  
Vol 70 (7) ◽  
pp. 1854-1872 ◽  
Author(s):  
Magdalena Bejger ◽  
Barbara Imiolczyk ◽  
Damien Clavel ◽  
Miroslaw Gilski ◽  
Agnieszka Pajak ◽  
...  
Keyword(s):  
Alkali Metal ◽  
Active Site ◽  
Metal Binding ◽  
Metal Ion ◽  
Catalytic Mechanism ◽  
Structural Data ◽  
Activation Loop ◽  
Plant Type ◽  
Substrate Preferences ◽  
Autocatalytic Cleavage

Plant-type L-asparaginases, which are a subclass of the Ntn-hydrolase family, are divided into potassium-dependent and potassium-independent enzymes with different substrate preferences. While the potassium-independent enzymes have already been well characterized, there are no structural data for any of the members of the potassium-dependent group to illuminate the intriguing dependence of their catalytic mechanism on alkali-metal cations. Here, three crystal structures of a potassium-dependent plant-type L-asparaginase fromPhaseolus vulgaris(PvAspG1) differing in the type of associated alkali metal ions (K+, Na+or both) are presented and the structural consequences of the different ions are correlated with the enzyme activity. As in all plant-type L-asparaginases, immature PvAspG1 is a homodimer of two protein chains, which both undergo autocatalytic cleavage to α and β subunits, thus creating the mature heterotetramer or dimer of heterodimers (αβ)2. The αβ subunits of PvAspG1 are folded similarly to the potassium-independent enzymes, with a sandwich of two β-sheets flanked on each side by a layer of helices. In addition to the `sodium loop' (here referred to as the `stabilization loop') known from potassium-independent plant-type asparaginases, the potassium-dependent PvAspG1 enzyme contains another alkali metal-binding loop (the `activation loop') in subunit α (residues Val111–Ser118). The active site of PvAspG1 is located between these two metal-binding loops and in the immediate neighbourhood of three residues, His117, Arg224 and Glu250, acting as a catalytic switch, which is a novel feature that is identified in plant-type L-asparaginases for the first time. A comparison of the three PvAspG1 structures demonstrates how the metal ion bound in the activation loop influences its conformation, setting the catalytic switch to ON (when K+is coordinated) or OFF (when Na+is coordinated) to respectively allow or prevent anchoring of the reaction substrate/product in the active site. Moreover, it is proposed that Ser118, the last residue of the activation loop, is involved in the potassium-dependence mechanism. The PvAspG1 structures are discussed in comparison with those of potassium-independent L-asparaginases (LlA, EcAIII and hASNase3) and those of other Ntn-hydrolases (AGA and Tas1), as well as in the light of noncrystallographic studies.

scholarly journals Elucidating the role of metal ions in carbonic anhydrase catalysis

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Jin Kyun Kim ◽  
Cheol Lee ◽  
Seon Woo Lim ◽  
Aniruddha Adhikari ◽  
Jacob T. Andring ◽  
...  
Keyword(s):  
Carbonic Anhydrase ◽  
Metal Ions ◽  
Metal Ion ◽  
Catalytic Mechanism ◽  
Chemical Properties ◽  
Water Network ◽  
Native Metal ◽  
Trigonal Bipyramidal ◽  
Electrostatic Effect

Abstract Why metalloenzymes often show dramatic changes in their catalytic activity when subjected to chemically similar but non-native metal substitutions is a long-standing puzzle. Here, we report on the catalytic roles of metal ions in a model metalloenzyme system, human carbonic anhydrase II (CA II). Through a comparative study on the intermediate states of the zinc-bound native CA II and non-native metal-substituted CA IIs, we demonstrate that the characteristic metal ion coordination geometries (tetrahedral for Zn2+, tetrahedral to octahedral conversion for Co2+, octahedral for Ni2+, and trigonal bipyramidal for Cu2+) directly modulate the catalytic efficacy. In addition, we reveal that the metal ions have a long-range (~10 Å) electrostatic effect on restructuring water network in the active site. Our study provides evidence that the metal ions in metalloenzymes have a crucial impact on the catalytic mechanism beyond their primary chemical properties.

Interaction of the adenoviral IVa2 protein with a truncated viral DNA packaging sequence

2009 ◽  
Vol 140 (1-3) ◽  
pp. 78-90 ◽  
Author(s):  
Teng-Chieh Yang ◽  
Qin Yang ◽  
Nasib Karl Maluf
Keyword(s):  
Dna Packaging ◽  
Viral Dna ◽  
Viral Dna Packaging ◽  
Packaging Sequence

scholarly journals A trapped human PPM1A–phosphopeptide complex reveals structural features critical for regulation of PPM protein phosphatase activity

2018 ◽  
Vol 293 (21) ◽  
pp. 7993-8008 ◽  
Author(s):  
Subrata Debnath ◽  
Dalibor Kosek ◽  
Harichandra D. Tagad ◽  
Stewart R. Durell ◽  
Daniel H. Appella ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Metal Ions ◽  
Active Site ◽  
Metal Binding ◽  
Metal Ion ◽  
Catalytic Domain ◽  
Protein Phosphatases ◽  
Random Order ◽  
Structural Features

Metal-dependent protein phosphatases (PPM) are evolutionarily unrelated to other serine/threonine protein phosphatases and are characterized by their requirement for supplementation with millimolar concentrations of Mg2+ or Mn2+ ions for activity in vitro. The crystal structure of human PPM1A (also known as PP2Cα), the first PPM structure determined, displays two tightly bound Mn2+ ions in the active site and a small subdomain, termed the Flap, located adjacent to the active site. Some recent crystal structures of bacterial or plant PPM phosphatases have disclosed two tightly bound metal ions and an additional third metal ion in the active site. Here, the crystal structure of the catalytic domain of human PPM1A, PPM1Acat, complexed with a cyclic phosphopeptide, c(MpSIpYVA), a cyclized variant of the activation loop of p38 MAPK (a physiological substrate of PPM1A), revealed three metal ions in the active site. The PPM1Acat D146E–c(MpSIpYVA) complex confirmed the presence of the anticipated third metal ion in the active site of metazoan PPM phosphatases. Biophysical and computational methods suggested that complex formation results in a slightly more compact solution conformation through reduced conformational flexibility of the Flap subdomain. We also observed that the position of the substrate in the active site allows solvent access to the labile third metal-binding site. Enzyme kinetics of PPM1Acat toward a phosphopeptide substrate supported a random-order, bi-substrate mechanism, with substantial interaction between the bound substrate and the labile metal ion. This work illuminates the structural and thermodynamic basis of an innate mechanism regulating the activity of PPM phosphatases.

Binding of metal ions and water molecules to nucleic acid bases: the influence of water molecule coordination to a metal ion on water–nucleic acid base hydrogen bonds

2019 ◽  
Vol 75 (3) ◽  
pp. 301-309
Author(s):  
Jelena M. Andrić ◽  
Ivana M. Stanković ◽  
Snežana D. Zarić
Keyword(s):  
Nucleic Acid ◽  
Hydrogen Bonds ◽  
Nucleic Acids ◽  
Metal Ions ◽  
Quantum Chemical Calculations ◽  
Quantum Chemical ◽  
Metal Ion ◽  
Water Molecules ◽  
Nucleic Acid Bases ◽  
Coordinated Water

The interactions of nucleic acid bases with non-coordinated and coordinated water molecules were studied by analyzing data in the Protein Data Bank (PDB) and by quantum chemical calculations. The analysis of the data in the crystal structures from the PDB indicates that hydrogen bonds involving oxygen or nitrogen atoms of nucleic acid bases and water molecules are shorter when water is bonded to a metal ion. These results are in agreement with the quantum chemical calculations on geometries and interaction energies of hydrogen bonds; the calculations on model systems show that hydrogen bonds of nucleic acid bases with water bonded to a metal ion are stronger than hydrogen bonds with non-coordinated water. These calculated values are similar to the strength of hydrogen bonds between nucleic acid bases. The results presented in this paper may be relevant to understand the role of water molecules and metal ions in the process of replication and stabilization of nucleic acids and also to understand the possible toxicity of metal ion interactions with nucleic acids.

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