MOTOR END PLATE REACTIVITY TO DIVALENT METAL IONS HISTOCHEMICAL STUDIES

Motor end plates in the tibialis anterior muscle of the rat were demonstrated by metal sulfide deposits following injection of aqueous solutions of lead, stannous, cadmium, zinc or cupric ions into the muscle in vivo or in vitro. The appearance of the end plates was similar to the structure demonstrated by cholinesterase staining, with visualization of the subneural apparatus. Neither metal binding nor cholinesterase activity was affected 4 weeks after dissection of the sciatic nerve, indicating that the metal binding site is postsynaptic. Freezing or formalin fixation of muscle prevented binding of all metal ions to the end plate without greatly affecting cholinesterase activity, indicating that these two activities of the end plate are distinct. Prior administration of acetylcholine, d-tubocurarine, neostigmine or diisopropyl fluorophosphate inhibited binding to the end plate of cadmium and zinc ions but did not alter binding of lead and stannous ions. By formation of a lake with alizarin red S previously injected in vivo intramuscularly, the release of calcium ions at the motor end plate following stimulation of the muscle through the nerve or administration of neostigmine was demonstrated. These results suggest a close relationship of the site of binding of divalent metal ions in the motor end plate to the site of calcium release, and a close but not identical relationship to the site of cholinesterase activity and the acetylcholine receptor.

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Regulation of divalent metal ions to the aggregation and membrane damage of human islet amyloid polypeptide oligomers

RSC Advances ◽

10.1039/d1ra00354b ◽

2021 ◽

Vol 11 (21) ◽

pp. 12815-12825

Author(s):

Yajie Wang ◽

Feihong Meng ◽

Tong Lu ◽

Chunyun Wang ◽

Fei Li

Keyword(s):

Metal Ions ◽

Metal Binding ◽

Membrane Damage ◽

Islet Amyloid Polypeptide ◽

Divalent Metal ◽

Human Islet ◽

Divalent Metal Ions ◽

Islet Amyloid ◽

Induced Membrane ◽

Human Islet Amyloid Polypeptide

Their is a counteraction between a decrease in the disruptive ability of metal-associated oligomer species and an increase in the quantity of oligomers promoted by the metal binding in the activity of hIAPP induced membrane damage.

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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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DEVELOPMENT OF THE NEUROMUSCULAR JUNCTION

The Journal of Cell Biology ◽

10.1083/jcb.55.1.93 ◽

1972 ◽

Vol 55 (1) ◽

pp. 93-103 ◽

Cited By ~ 24

Author(s):

Thomas L. Lentz

Keyword(s):

Schwann Cell ◽

Cholinesterase Activity ◽

Neuromuscular Junctions ◽

Synaptic Cleft ◽

Axon Terminals ◽

End Plate ◽

Motor End Plate ◽

Motor End Plates

To determine the effects of nerve explants on the integrity of motor end plates in vitro, cholinesterase activity and structure of end plates were compared in newt muscle denervated in vivo, cultured in the absence of nerve explants, and cultured in the presence of sensory ganglia. In neuromuscular junctions denervated in vivo or in vitro, the synaptic vesicles become clumped and fragmented. A few intact vesicles escape into the synaptic cleft. Axon terminals degenerate until they are left as residual bodies within the Schwann cell cytoplasm. Junctional folds on the muscle surface are reduced in height and are no longer evident once traces of axoplasm within the Schwann cell disappear. End plate cholinesterase activity is reduced as junctional folds are lost. When muscle is cultured in the presence of a sensory ganglion, the terminal axoplasm degenerates in the same manner but junctional folds persist on the muscle surface. Moderately intense cholinesterase activity remains in association with the junctional folds, so that normal motor end plates are maintained in the absence of innervation. These results show that degenerative changes in the structure of the motor end plate and loss of cholinesterase activity occurring in organ culture as a result of denervation can be retarded by nerve explants that do not directly innervate the muscle.

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Interactions of DNA with divalent metal ions. III. Extent of metal binding: Experiment and theory

Biopolymers ◽

10.1002/bip.360210117 ◽

1982 ◽

Vol 21 (1) ◽

pp. 219-232 ◽

Cited By ~ 31

Author(s):

Joseph Granot ◽

David R. Kearns

Keyword(s):

Metal Ions ◽

Metal Binding ◽

Divalent Metal ◽

Divalent Metal Ions ◽

Binding Experiment

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Influence of monovalent metal ions on metal binding and catalytic activity of the 10–23 DNAzyme

Biological Chemistry ◽

10.1515/hsz-2020-0207 ◽

2020 ◽

Vol 402 (1) ◽

pp. 99-111

Author(s):

Hannah Rosenbach ◽

Jan Borggräfe ◽

Julian Victor ◽

Christine Wuebben ◽

Olav Schiemann ◽

...

Keyword(s):

Metal Ions ◽

Metal Binding ◽

Divalent Cations ◽

Reaction Rates ◽

Divalent Metal ◽

Divalent Metal Ions ◽

Monovalent Metal ◽

Biophysical Techniques ◽

Target Rna

AbstractDeoxyribozymes (DNAzymes) are single-stranded DNA molecules that catalyze a broad range of chemical reactions. The 10–23 DNAzyme catalyzes the cleavage of RNA strands and can be designed to cleave essentially any target RNA, which makes it particularly interesting for therapeutic and biosensing applications. The activity of this DNAzyme in vitro is considerably higher than in cells, which was suggested to be a result of the low intracellular concentration of bioavailable divalent cations. While the interaction of the 10–23 DNAzyme with divalent metal ions was studied extensively, the influence of monovalent metal ions on its activity remains poorly understood. Here, we characterize the influence of monovalent and divalent cations on the 10–23 DNAzyme utilizing functional and biophysical techniques. Our results show that Na+ and K+ affect the binding of divalent metal ions to the DNAzyme:RNA complex and considerably modulate the reaction rates of RNA cleavage. We observe an opposite effect of high levels of Na+ and K+ concentrations on Mg2+- and Mn2+-induced reactions, revealing a different interplay of these metals in catalysis. Based on these findings, we propose a model for the interaction of metal ions with the DNAzyme:RNA complex.

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