Exploring the dynamic nature of divalent metal ions involved in DNA cleavage by CRISPR-Cas12a

CRISPR-Cas12a has been widely used in genome editing and nucleic acid detection. In both of these applications, Cas12a cleaves target DNA in a divalent metal ion-dependent manner. However, when and...

Influence of divalent metal ion (Zn2+) on Magnetic properties, Curie temperature and DC Electrical properties in Ce3+ Substituted Ni-Zn Ferrites.

Nanoparticles of Ni1-xZnxCe0.1Fe1.9O4 ferrite substituted with Zn2+ ion concentrations (0.0, 0.2, 0.4, 0.6, 0.8 & 1.0) have been synthesised via aqueous citrate precursor auto combustion method. The obtained Powder X-ray diffraction data suggests a good crystalline phase; the crystallite size exists in the range of 14~38 nm. The lattice constant of samples increases with Zn2+ concentration. It validates Vegard's law and the decreasing trends in porosity of the samples are identified. The inhomogeneity in the grains was confirmed from FE-SEM. The EDS spectrogram attributes the good stoichiometry in the product. The Thermogravimetric analysis reveals that the crystallization occurred within the temperature 800˚C. The high-temperature DC conductivity of the samples shows that NTC behaviour. The Curie temperature and activation energy are estimated and the activation energy of carriers are found to be more at the paramagnetic region. The magnetic saturation ('Ms*') has maximum for Zn2+ = 0.2 (57.7 emu/g) and coercive field (Hc) related with a radius of occupancy of Zn2+ ion validates the relation Hc∝1/r and also, the value of the remanence ratio suggests that isotropic magnetization took place in the heterogeneous material.

Magnetic and Structural Behaviour of Cobalt & Nickel Substituted Calcium W-Type Nanosize Hexaferrites

The available literature and research work on W-type hexaferrites is mainly focused on Co- and Zn-based calcium W-type hexagonal ferrites with a variety of cationic substitutions. The Modifications in the properties of the Calcium W-type ferrite based on Ni2+ as the divalent metal ion, however, is not studied sufficiently in the research literature vailable. In this study, the focus is mainly on the effects of substitution of Ni2+ on the properties of CaCo2W exaferrites. The investigations carried out are mainly XRD, SEM and VSM. The main objective of this research investigation is to study the effect of substitution of Nickel and Cobalt on the structural and magnetic properties of calcium W-type hexaferrite CaCo2-xNixFe16O27 (x=0, 1 and 2). XRD analysis and characterization revealed slight decrease in the values of lattice constants ‘a’ and ‘c’ with increase in concentration ‘x’. The particle size was confirmed from SEM and TEM images. The analysis of VSM for magnetic properties reveals decrease in coercivity and increase in the values of saturation magnetization as concentration increases. The results of measurements made bythe various experimental techniques and the observations were compared to understand the crystalline and magnetic structure of the compounds

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.

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

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.

H101G Mutation in Rat Lens αB-Crystallin Alters Chaperone Activity and Divalent Metal Ion Binding

Background: The molecular chaperone function of αB-crystallins is heavily involved in maintaining lens transparency and the development of cataracts. Objective: To study whether divalent metal ion binding improves the stability and αB-crystallin chaperone activity. Results: Substitution of His101 with glycine resulted in structural and functional changes. Spectral analysis and chaperone-like activity assays showed that substitution of glycine resulted in a higher percentage of random coils, increased hydrophobicity, and 22±2% higher chaperone-like activity. Whereas in the presence of the Cu2+ ion, H101G exhibited 32±1% less chaperone-like activity compared to the wild type. Conclusion: Cu2+ has been reported to enhance the chaperone-like activity of lens α-crystallin. Our results indicate that H101 is the predominant Cu2binding site, and the mutation resulted in a partial unfolding that impaired the binding of Cu2+ to H101 residue. In conclusion, this study further helps to understand the important binding site for Cu2+ to αB-crystallin.

A metal ion orients SARS-CoV-2 mRNA to ensure accurate 2′-O methylation of its first nucleotide

The SARS-CoV-2 nsp16/nsp10 enzyme complex modifies the 2′-OH of the first transcribed nucleotide of the viral mRNA by covalently attaching a methyl group to it. The 2′-O methylation of the first nucleotide converts the status of mRNA cap from Cap-0 to Cap-1, and thus, helps the virus evade immune surveillance in host cells. Here, we report two structures of nsp16/nsp10 representing pre- and post-release states of the RNA product (Cap-1). We observe overall widening of the enzyme upon product formation, and an inward twisting motion in the substrate binding region upon product release. These conformational changes reset the enzyme for the next round of catalysis. The structures also identify a unique binding mode and the importance of a divalent metal ion for 2′-O methylation. We also describe underlying structural basis for the perturbed enzymatic activity of a clinical variant of SARS-CoV-2, and a previous SARS-CoV outbreak strain.