Structure of New Binary and Ternary DNA Polymerase Complexes From Bacteriophage RB69

DNA polymerase plays a critical role in passing the genetic information of any living organism to its offspring. DNA polymerase from enterobacteria phage RB69 (RB69pol) has both polymerization and exonuclease activities and has been extensively studied as a model system for B-family DNA polymerases. Many binary and ternary complex structures of RB69pol are known, and they all contain a single polymerase-primer/template (P/T) DNA complex. Here, we report a crystal structure of the exonuclease-deficient RB69pol with the P/T duplex in a dimeric form at a resolution of 2.2 Å. The structure includes one new closed ternary complex with a single divalent metal ion bound and one new open binary complex in the pre-insertion state with a vacant dNTP-binding pocket. These complexes suggest that initial binding of the correct dNTP in the open state is much weaker than expected and that initial binding of the second divalent metal ion in the closed state is also much weaker than measured. Additional conformational changes are required to convert these complexes to high-affinity states. Thus, the measured affinities for the correct incoming dNTP and divalent metal ions are average values from many conformationally distinctive states. Our structure provides new insights into the order of the complex assembly involving two divalent metal ions. The biological relevance of specific interactions observed between one RB69pol and the P/T duplex bound to the second RB69pol observed within this dimeric complex is discussed.

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Divalent metal ion-mediated assembly of spherical nucleic acids: the case study of Cu2+

Physical Chemistry Chemical Physics ◽

10.1039/c5cp05202e ◽

2015 ◽

Vol 17 (45) ◽

pp. 30292-30299 ◽

Cited By ~ 4

Author(s):

Jang Ho Joo ◽

Jae-Seung Lee

Keyword(s):

Nucleic Acids ◽

Metal Ions ◽

Metal Ion ◽

Divalent Metal ◽

Divalent Metal Ions ◽

New Strategy ◽

Divalent Metal Ion ◽

Spherical Nucleic Acids

A new strategy for reversibly assembling spherical nucleic acids (SNAs) is demonstrated based on the coordinative binding of divalent metal ions, particularly Cu2+, to nucleobases.

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Effect of divalent metal ions on annexin-mediated aggregation of asolectin liposomes.

Acta Biochimica Polonica ◽

10.18388/abp.2000_3988 ◽

2000 ◽

Vol 47 (3) ◽

pp. 675-683 ◽

Cited By ~ 1

Author(s):

V I Mel'gunov ◽

E I Akimova ◽

K S Krasavchenko

Keyword(s):

Metal Ions ◽

Metal Ion ◽

Relative Concentration ◽

Toxic Metal ◽

Transition Metal Ions ◽

Divalent Metal ◽

Divalent Metal Ions ◽

Large Unilamellar Vesicles ◽

Protein Molecules ◽

Divalent Metal Ion

Annexins belong to a family of Ca2+- and phospholipid-binding proteins that can mediate the aggregation of granules and vesicles in the presence of Ca2+. We have studied the effects of different divalent metal ions on annexin-mediated aggregation of liposomes using annexins isolated from rabbit liver and large unilamellar vesicles prepared from soybean asolectin II-S. In the course of these studies, we have found that annexin-mediated aggregation of liposomes can be driven by various earth and transition metal ions other than Ca2+. The ability of metal ions to induce annexin-mediated aggregation decreases in the order: Cd2+ > Ba2+, Sr2+ > Ca2+ > Mn2+ > Ni2+ > Co2+. Annexin-mediated aggregation of vesicles is more selective to metal ions than the binding of annexins to membranes. We speculate that not every type of divalent metal ion can induce conformational change sufficient to promote the interaction of annexins either with two opposing membranes or with opposing protein molecules. Relative concentration ratios of metal ions in the intimate environment may be crucial for the functioning of annexins within specialized tissues and after treatment with toxic metal ions.

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Structural and thermodynamic insights into a novel Mg2+–citrate-binding protein from the ABC transporter superfamily

Acta Crystallographica Section D Structural Biology ◽

10.1107/s2059798321010457 ◽

2021 ◽

Vol 77 (12) ◽

Author(s):

Suraj Kumar Mandal ◽

Shankar Prasad Kanaujia

Keyword(s):

Metal Ions ◽

Abc Transporter ◽

Metal Ion ◽

Binding Protein ◽

Substrate Binding ◽

Divalent Metal ◽

Uptake System ◽

Divalent Metal Ions ◽

Gram Negative ◽

Divalent Metal Ion

More than one third of proteins require metal ions to accomplish their functions, making them obligatory for the growth and survival of microorganisms in varying environmental niches. In prokaryotes, besides their involvement in various cellular and physiological processes, metal ions stimulate the uptake of citrate molecules. Citrate is a source of carbon and energy and is reported to be transported by secondary transporters. In Gram-positive bacteria, citrate molecules are transported in complex with divalent metal ions, whereas in Gram-negative bacteria they are translocated by Na+/citrate symporters. In this study, the presence of a novel divalent-metal-ion-complexed citrate-uptake system that belongs to the primary active ABC transporter superfamily is reported. For uptake, the metal-ion-complexed citrate molecules are sequestered by substrate-binding proteins (SBPs) and transferred to transmembrane domains for their transport. This study reports crystal structures of an Mg2+–citrate-binding protein (MctA) from the Gram-negative thermophilic bacterium Thermus thermophilus HB8 in both apo and holo forms in the resolution range 1.63–2.50 Å. Despite binding various divalent metal ions, MctA possesses the coordination geometry to bind its physiological metal ion, Mg2+. The results also suggest an extended subclassification of cluster D SBPs, which are known to bind and transport divalent-metal-ion-complexed citrate molecules. Comparative assessment of the open and closed conformations of the wild-type and mutant MctA proteins suggests a gating mechanism of ligand entry following an `asymmetric domain movement' of the N-terminal domain for substrate binding.

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Transfer of divalent metal ions into the organic phase of the systems water–chloroform–ZnCl2–CdCl2–HgCl2–NaOH–(NaCl)–n-alkyltri(oxyethylene)carboxylic acid

Canadian Journal of Chemistry ◽

10.1139/v88-414 ◽

1988 ◽

Vol 66 (10) ◽

pp. 2640-2646 ◽

Cited By ~ 2

Author(s):

Jerzy Strzelbicki ◽

Witold Charewicz ◽

Jorg Beger ◽

Lutz Hinz

Keyword(s):

Carboxylic Acid ◽

Aqueous Solutions ◽

Metal Ions ◽

Organic Phase ◽

Metal Ion ◽

Alkyl Chain ◽

Divalent Metal ◽

Divalent Metal Ions ◽

Chemical Equilibria ◽

Divalent Metal Ion

The influence of the length of an n-alkyl chain incorporated into the molecule of an n-alkyltri(oxyethylene) carboxylic acid of a general formula CnH(2n+1)(OCH2CH2)3OCH2CO2H(n = 6, 8, 10, and 12, respectively) on transfer of the zinc group element ions into chloroform was investigated. Metal ions were transferred from the aqueous solutions of ZnCl2, CdCl2, and HgCl2 at initial concentrations of 0.835 mM and 1.67 mM with, and without, added NaCl. From the aqueous solutions to which no NaCl was added, the selectivity of transfer of divalent metal ions into the organic phase was low.The selectivity of transfer of the divalent metal ions improved remarkably when NaCl was added to the aqueous phase due to the chemical equilibria involving complexation of the divalent metal ions by Cl−, OH−, and n-alkyltri(oxyethylene) carboxylic acid. In most systems (with NaCl added), Zn(II) was separated from Cd(II) and Hg(II). Less lipophilic n-alkyltri(oxyethylene) carboxylic acids (with a shorter n-alkyl chain incorporated) exhibited higher selectivity and efficiency as divalent metal ion carriers.

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HK97 gp74 Possesses an α-Helical Insertion in the ββα Fold That Affects Its Metal Binding, cos Site Digestion, and In Vivo Activities

Journal of Bacteriology ◽

10.1128/jb.00644-19 ◽

2020 ◽

Vol 202 (8) ◽

Author(s):

Sasha A. Weiditch ◽

Sarah C. Bickers ◽

Diane Bona ◽

Karen L. Maxwell ◽

Voula Kanelis

Keyword(s):

Metal Binding ◽

Metal Ion ◽

Divalent Metal ◽

Metal Ion Binding ◽

Divalent Metal Ions ◽

Nmr Studies ◽

Phage Dna ◽

Divalent Metal Ion ◽

Phage Morphogenesis

ABSTRACT The last gene in the genome of the bacteriophage HK97 encodes gp74, an HNH endonuclease. HNH motifs contain two conserved His residues and an invariant Asn residue, and they adopt a ββα structure. gp74 is essential for phage head morphogenesis, likely because gp74 enhances the specific endonuclease activity of the HK97 terminase complex. Notably, the ability of gp74 to enhance the terminase-mediated cleavage of the phage cos site requires an intact HNH motif in gp74. Mutation of H82, the conserved metal-binding His residue in the HNH motif, to Ala abrogates gp74-mediated stimulation of terminase activity. Here, we present nuclear magnetic resonance (NMR) studies demonstrating that gp74 contains an α-helical insertion in the Ω-loop, which connects the two β-strands of the ββα fold, and a disordered C-terminal tail. NMR data indicate that the Ω-loop insert makes contacts to the ββα fold and influences the ability of gp74 to bind divalent metal ions. Further, the Ω-loop insert and C-terminal tail contribute to gp74-mediated DNA digestion and to gp74 activity in phage morphogenesis. The data presented here enrich our molecular-level understanding of how HNH endonucleases enhance terminase-mediated digestion of the cos site and contribute to the phage replication cycle. IMPORTANCE This study demonstrates that residues outside the canonical ββα fold, namely, the Ω-loop α-helical insert and a disordered C-terminal tail, regulate the activity of the HNH endonuclease gp74. The increased divalent metal ion binding when the Ω-loop insert is removed compared to reduced cos site digestion and phage formation indicates that the Ω-loop insert plays multiple regulatory roles. The data presented here provide insights into the molecular basis of the involvement of HNH proteins in phage DNA packing.

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Activation of Bovine Factor X in the Presence of Calcium, Magnesium, Barium or Manganese ion

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648669 ◽

1978 ◽

Vol 40 (02) ◽

pp. 358-367 ◽

Cited By ~ 2

Author(s):

Robert H Yue ◽

Menard M Gertler

Keyword(s):

Metal Ions ◽

Binding Site ◽

Metal Ion ◽

Factor Xa ◽

Divalent Metal ◽

Activation Process ◽

Factor X ◽

Divalent Metal Ion ◽

Russell’S Viper ◽

Bovine Factor

SummaryThe binding of divalent metal ions to bovine factor X, factor Xa and the coagulant protein in Russell’s viper venom was studied by the technique of fluorescence quenching. Titration of factor X with Ca+2, Mg+2 or Ba+2 revealed that these metal ions can bind to factor X. A tightly binding site(s) was observed with Kd of 79 and 98 μM for Ca+2 and Mg+2 respectively. A loosely binding site(s) was evident with Kd of 0.55, 0.50 and 0.35 mM for Ca+2, Mg+2 and Ba+2 respectively. The quenching phenomenon was also observed when Mn+2 was used as titrant but factor X precipitated out when the concentration of Mn+2 was 10 mM. The binding of Ca+2, Mg+2, Ba+2 or Mn+2 to bovine factor Xa or to the purified coagulant fraction of Russell’s viper venom was very weak in each case.In the absence of Ca+2, the coagulation fraction of Russell’s viper venom could not activate bovine factor X. Activation of factor X was achieved when Ca+2 was replaced by either Mg+2, Ba+2 or Mn+2. When the concentration of these ions were 5 mM, the efficiency of factor Xa generation was estimated to be: Ca+2> Mg+2> Ba+2> Mn+2. Higher concentration of Mg+2, Ba+2, or Mn+2 retarded the activation process. However, Ca+2, Mg+2, Ba+2 or Mn+2 has little or no influence on the esterase activity of factor Xa or purified Rusell’s viper venom.The results suggest that complexation of divalent metal ion with factor X is prerequisite in the activation process. The binding of Mg+2, Ba+2 or Mn+2 to these loosely binding sites might have altered the geometrical configuration as well as the electrostatic environment on factor X significantly. Thus, it is more difficult to form the binary complex and a slower generation of factor Xa results. Therefore, divalent metal ion serves as a dual role in the activation of factor X to factor Xa depending upon the ionic concentration.

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Modeling of neotame and fructose thermochemistry: Comparison with mono and divalent metal ions by Computational and experimental approach

Scientific Reports ◽

10.1038/s41598-019-54626-9 ◽

2019 ◽

Vol 9 (1) ◽

Author(s):

Deepali Sharma ◽

Suvardhan Kanchi ◽

Ayyappa Bathinapatla ◽

Inamuddin ◽

Abdullah M. Asiri

Keyword(s):

Metal Ions ◽

Metal Ion ◽

Density Functional ◽

Divalent Metal ◽

Basis Set ◽

Hydroxyl Groups ◽

Specific Cell ◽

Water Medium ◽

Artificial Sweetener ◽

Divalent Metal Ions

AbstractThe metal complexes can demonstrate various interesting biological activities in the human body. However, the role of certain metal ions for specific cell activities is still subject to debate. This study is aimed at comparing the thermochemical properties of neotame (artificial sweetener) and α, β-fructose in gas phase and water medium. The interaction of α and β-fructose, neotame with monovalent and divalent metal ions was studied and comprehended by density functional theory (DFT) using B3LYP functional, 6–311 + G (d, p) and D3 basis set. Metal ion affinities (MIA) values depicted that ionic radius of metal ions played an important role in the interaction of α, β-fructose and neotame. The ∆G parameter was calculated to predict and understand the interaction of metal ions with α and β-fructose, neotame. The results suggested that the presence of hydroxyl groups and oxygen atoms in sugar molecules acted as preferred sites for the binding and interaction of mono and divalent ions. For the first time computational study has been introduced in the present study to review the progress in the application of metal binding with sugar molecules especially with neotame. Moreover, voltammetric behaviour of neotame-Zn2+ was studied using cyclic and differential pulse voltammetry. The obtained results suggest that the peak at −1.13 V is due to the reduction of Zn2+ in 0.1 M phosphate buffer medium at pH 5.5. Whereas, addition of 6-fold higher concentration of neotame to the ZnCl2.2H2O resulted in a new irreversible cathodic peak at −0.83, due to the reduction of neotame-Zn2+ complex. The Fourier transform infrared spectroscopy (FTIR) results indicates that the β-amino group (-NH) and carboxyl carbonyl (-C = O) groups of neotame is participating in the chelation process, which is further supported by DFT studies. The findings of this study identify the efficient chelation factors as major contributors into metal ion affinities, with promising possibilities to determine important biological processes in cell wall and glucose transmembrane transport.

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Thermoelectric properties of Li-doped Cu0.95-xM0.05LixO (M=Mn, Ni, Zn)

MRS Proceedings ◽

10.1557/opl.2012.1571 ◽

2012 ◽

Vol 1490 ◽

pp. 69-73

Author(s):

N. Yoshida ◽

T. Naito ◽

H. Fujishiro

Keyword(s):

Thermal Conductivity ◽

Electrical Resistivity ◽

Thermoelectric Properties ◽

Metal Ion ◽

Figure Of Merit ◽

Divalent Metal ◽

Thermoelectric Figure Of Merit ◽

Divalent Metal Ions ◽

Thermoelectric Figure ◽

Divalent Metal Ion

ABSTRACTThermoelectric properties of the Li-doped Cu0.95-xM0.05LixO (M=divalent metal ion; Mn, Ni, Zn) were investigated at the temperature up to 1273 K. In the doped divalent metal ions, Zn2+ ion was the most effective to reduce the thermal conductivity, and the Ni2+ substitution was preferable to decrease the electrical resistivity. For the Cu0.95-xNi0.05LixO sample at x=0.03, the maxima of the dimensionless thermoelectric figure of merit ZT and the power factor P at 1246 K were 4.2×10-2 and 1.6 ×10-4 W/K2m, respectively. The enhancement of the thermoelectric properties of the Li-doped Cu0.95-xM0.05LixO system was discussed.

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