In Vivo Metal Ion Imaging Using Fluorescent Sensors

2016 ◽  
pp. 139-152
Author(s):  
Genevieve C. Van de Bittner ◽  
Tasuku Hirayama
Keyword(s):  
Metal Ion ◽  
Fluorescent Sensors ◽  
Ion Imaging

scholarly journals Crystallographic and X-ray absorption spectroscopic characterization of Helicobacter pylori UreE bound to Ni2+ and Zn2+ reveals a role for the disordered C-terminal arm in metal trafficking

2012 ◽  
Vol 441 (3) ◽  
pp. 1017-1035 ◽  
Author(s):  
Katarzyna Banaszak ◽  
Vlad Martin-Diaconescu ◽  
Matteo Bellucci ◽  
Barbara Zambelli ◽  
Wojciech Rypniewski ◽  
...  
Keyword(s):  
Helicobacter Pylori ◽  
Metal Ions ◽  
Metal Binding ◽  
Metal Ion ◽  
Mutual Influence ◽  
Accurate Information ◽  
Metal Ion Binding ◽  
X Ray ◽  
X Ray Absorption

The survival and growth of the pathogen Helicobacter pylori in the gastric acidic environment is ensured by the activity of urease, an enzyme containing two essential Ni2+ ions in the active site. The metallo-chaperone UreE facilitates in vivo Ni2+ insertion into the apoenzyme. Crystals of apo-HpUreE (H. pylori UreE) and its Ni2+- and Zn2+-bound forms were obtained from protein solutions in the absence and presence of the metal ions. The crystal structures of the homodimeric protein, determined at 2.00 Å (apo), 1.59 Å (Ni2+) and 2.52 Å (Zn2+) resolution, show the conserved proximal and solvent-exposed His102 residues from two adjacent monomers invariably involved in metal binding. The C-terminal regions of the apoprotein are disordered in the crystal, but acquire significant ordering in the presence of the metal ions due to the binding of His152. The analysis of X-ray absorption spectral data obtained using solutions of Ni2+- and Zn2+-bound HpUreE provided accurate information of the metal-ion environment in the absence of solid-state effects. These results reveal the role of the histidine residues at the protein C-terminus in metal-ion binding, and the mutual influence of protein framework and metal-ion stereo-electronic properties in establishing co-ordination number and geometry leading to metal selectivity.

In vivo blood metal ion levels in patients after total shoulder arthroplasty

2019 ◽  
Vol 28 (3) ◽  
pp. 539-546 ◽  
Author(s):  
Tobias Reiner ◽  
Nina Bader ◽  
Benjamin Panzram ◽  
Matthias Bülhoff ◽  
Georg Omlor ◽  
...  
Keyword(s):  
Shoulder Arthroplasty ◽  
Metal Ion ◽  
Total Shoulder Arthroplasty

In Vivo Evaluation of Micronucleus Frequencies in Buccal Mucosal Cells of Orthodontic Patients with and Without Fluoride Use

2021 ◽  
pp. 030157422110373
Author(s):  
Prasad Chitra ◽  
GS Prashantha ◽  
Arun Rao ◽  
Harshvardhan S Jois
Keyword(s):  
Metal Ion ◽  
Orthodontic Treatment ◽  
Biological Tissues ◽  
Control Group ◽  
Time Interval ◽  
Ion Release ◽  
In Vivo Evaluation ◽  
Metal Ion Release ◽  
Mucosal Cells

Introduction: Fluoride agents to prevent white spot lesions are used often during orthodontic treatment. The beneficial effects of fluoride, when consumed within permissible limits on dental structures, are well known. Their implications on underlying biological tissues, however, are unknown. Mouthwashes and dentifrices with fluorides are associated with metal ion release into the mouth with possible cell genotoxicity. Since these cariostatic agents are frequently used during orthodontic therapy, a deeper understanding of the effects of fluoride on oral tissues was considered necessary. Methodology: Three groups of patients (30 each)—group 1 (untreated controls), group 2 (non-fluoridated), and group 3 (Fluoridated) were analyzed. Patients in groups 2 and 3 were bonded with the same bracket prescription and treated with similar archwire sequences. Buccal mucosal cells at 4 specific time periods (before treatment, 1 week, 30 days, and 6 months) were collected, using a wooden tongue depressor, and assessed for any nuclear abnormalities. Comparisons of changes were made with an untreated control group and also between the non-fluoridated and fluoridated groups. Relevant conclusions were drawn after analysis of the results. Results: Greater number of nuclei were observed at the 30-day time interval in the fluoridated group, which was statistically significant at P < .001. Conclusion: Use of fluoridated oral hygiene products in patients undergoing fixed orthodontic treatment with NiTi archwires could increase the risk of micronuclei formation in buccal mucosal cells.

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

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.

scholarly journals SYNTHESIS, MOLLUSCICIDALAND ANTIMICROBIAL POTENTIALITIES OF IRON TRIAD MONONUCLEAR METAL COMPLEXES INCORPORATING TRIDENTATE ASYMMETRICAL SCHIFF BASE LIGANDS CONTAINING SOFT SULFUR COORDINATING ATOM

2019 ◽  
pp. 103-113
Author(s):  
AMAL M ALOSAIMI ◽  
INES EL MANNOUBI ◽  
SAMI A ZABIN
Keyword(s):  
Schiff Base ◽  
Metal Complexes ◽  
Metal Ion ◽  
Lethal Dose ◽  
Analytical Techniques ◽  
Central Metal ◽  
Schiff Base Ligands ◽  
Molluscicidal Activity

Objective: This work aimed at synthesizing tridentates asymmetrical Schiff base ligands containing sulfur atom and using them for preparing metal complexes with the iron triad metals. The prepared compounds were assayed in vitro for antimicrobial potential and in vivo molluscicidal activity. Methods: The unsymmetrical tridentate Schiff bases (SL1, SL2, and SL3) were prepared using 2-aminothiophenol as primary amine and condensed with 2-carboxybenzaldehyde, 2-hydroxy-1-naphthaldehyde, and 7-formyl-8-hydroxyquinoline. These ligands were used in preparing metal complexes with iron triad metals. The synthesized Schiff base ligands and their corresponding metal complexes were characterized and their proposed structures were confirmed using different physical and spectroscopic analytical techniques. All ligands and their corresponding metal complexes were assayed against different bacterial and fungal strains using the agar disk-diffusion technique. The molluscicidal activity was performed according to the standard reported methods as cited in the literature and by observing the toxicity and lethal dose according to the WHO guidelines. Results: The synthesized ligands behave as tridentate (NOS) ligands and form mononuclear complexes with the general formula [M(SL)2] with an octahedral geometry around the central metal ion. Metal complexes were non-electrolytic in nature. The in vitro antibacterial and antifungal examination results showed weak activity of the ligands, and there was enhanced activity with the complexes. The in vivo molluscicidal activity of the tested compounds showed good activity. Conclusion: The targeted compounds were prepared successfully, characterized, and showed some biological activity but lower than the standard reference drugs.

scholarly journals A high-speed, bright, red fluorescent voltage sensor to detect neural activity

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Connor Beck ◽  
Yiyang Gong
Keyword(s):  
Blue Light ◽  
Neural Activity ◽  
High Speed ◽  
Fluorescent Protein ◽  
Resonance Energy ◽  
Fluorescent Sensors ◽  
Fast Kinetics ◽  
Spectral Separation ◽  
Green Fluorescent

Abstract Genetically encoded voltage indicators (GEVIs) have emerged as a technology to optically record neural activity with genetic specificity and millisecond-scale temporal resolution using fluorescence microscopy. GEVIs have demonstrated ultra-fast kinetics and high spike detection fidelity in vivo, but existing red-fluorescent voltage indicators fall short of the response and brightness achieved by green fluorescent protein-based sensors. Furthermore, red-fluorescent GEVIs suffer from incomplete spectral separation from green sensors and blue-light-activated optogenetic actuators. We have developed Ace-mScarlet, a red fluorescent GEVI that fuses Ace2N, a voltage-sensitive inhibitory rhodopsin, with mScarlet, a bright red fluorescent protein (FP). Through fluorescence resonance energy transfer (FRET), our sensor detects changes in membrane voltage with high sensitivity and brightness and has kinetics comparable to the fastest green fluorescent sensors. Ace-mScarlet’s red-shifted absorption and emission spectra facilitate virtually complete spectral separation when used in combination with green-fluorescent sensors or with blue-light-sensitive sensors and rhodopsins. This spectral separation enables both simultaneous imaging in two separate wavelength channels and high-fidelity voltage recordings during simultaneous optogenetic perturbation.

scholarly journals Antibodies against Thrombospondin-Related Anonymous Protein Do Not Inhibit Plasmodium Sporozoite Infectivity In Vivo

2000 ◽  
Vol 68 (6) ◽  
pp. 3667-3673 ◽  
Author(s):  
Soren Gantt ◽  
Cathrine Persson ◽  
Keith Rose ◽  
Ashley J. Birkett ◽  
Ruben Abagyan ◽  
...  
Keyword(s):  
Metal Ion ◽  
Competitive Inhibition ◽  
Polyclonal Antibodies ◽  
Three Dimensional ◽  
Calcium Ionophore ◽  
Plasmodium Yoelii ◽  
Gliding Motility ◽  
Vaccine Antigen ◽  
Dimensional Structure

ABSTRACT Thrombospondin-related anonymous protein (TRAP), a candidate malaria vaccine antigen, is required for Plasmodiumsporozoite gliding motility and cell invasion. For the first time, the ability of antibodies against TRAP to inhibit sporozoite infectivity in vivo is evaluated in detail. TRAP contains an A-domain, a well-characterized adhesive motif found in integrins. We modeled here a three-dimensional structure of the TRAP A-domain of Plasmodium yoelii and located regions surrounding the MIDAS (metal ion-dependent adhesion site), the presumed business end of the domain. Mice were immunized with constructs containing these A-domain regions but were not protected from sporozoite challenge. Furthermore, monoclonal and rabbit polyclonal antibodies against the A-domain, the conserved N terminus, and the repeat region of TRAP had no effect on the gliding motility or sporozoite infectivity to mice. TRAP is located in micronemes, secretory organelles of apicomplexan parasites. Accordingly, the antibodies tested here stained cytoplasmic TRAP brightly by immunofluorescence. However, very little TRAP could be detected on the surface of sporozoites. In contrast, a dramatic relocalization of TRAP onto the parasite surface occurred when sporozoites were treated with calcium ionophore. This likely mimics the release of TRAP from micronemes when a sporozoite contacts its target cell in vivo. Contact with hepatoma cells in culture also appeared to induce the release of TRAP onto the surface of sporozoites. If large amounts of TRAP are released in close proximity to its cellular receptor(s), effective competitive inhibition by antibodies may be difficult to achieve.

An immunochemical study of Neurospora nucleases

1986 ◽  
Vol 64 (2) ◽  
pp. 106-116 ◽  
Author(s):  
Murray J. Fraser ◽  
Terry Y.-K. Chow ◽  
Helga Cohen ◽  
Helena Koa
Keyword(s):  
Metal Ion ◽  
Sodium Dodecyl ◽  
Limited Proteolysis ◽  
Mung Bean Nuclease ◽  
S1 Nuclease ◽  
Immunochemical Study ◽  
Dna And Rna ◽  
Cell Compartments ◽  
Divalent Metal Ion

Nucleases derived from Neurospora crassa mycelia with neutral single-strand (ss) endodeoxyribonuclease activity have been examined by immunochemical techniques and by sodium dodecyl sulfate – DNA gel electrophoresis. All of the intracellular nucleases, which have different divalent metal ion requirements, different strand specificities with single- and double-strand DNA, different modes of action on DNA and RNA, and other distinguishing characteristics, are immunochemically related to Neurospora endo-exonuclease. The evidence indicates that these enzymes are derived from one or more related large, inactive (precursor?) polypeptides that are first converted to 75- to 80-kdalton active polypeptide(s) which are very protease sensitive. Further limited proteolysis results in the production of the various active forms of nuclease studied here. Some proteolytic conversions may occur in a controlled manner in vivo in different cell compartments, but others are very likely artifacts resulting from uncontrolled proteolysis during extraction and isolation. The intracellular forms of Neurospora endo-exonuclease are immunologically cross-active with ss-DNA-binding nucleases isolated from Aspergillus nidulans and Saccharomyces cerevisiae. They are not immunochemically related to two extracellular Neurospora nucleases, the pancreatic DNase-I-like DNase A and a ss-specific exonuclease, and they are also not related to other fungal and plant nucleases with ss-specific endonuclease activity such as the S1 nuclease of Aspergillus oryzae, the P1 nuclease of Penicillium citrinum, and mung bean nuclease.

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