scholarly journals Design of a new DNA-polyintercalating drug, a bisacridinyl peptidic analogue of Triostin A

1985 ◽  
Vol 225 (3) ◽  
pp. 829-832 ◽  
Author(s):  
N Helbecque ◽  
J L Bernier ◽  
J P Hénichart
Keyword(s):  
Thermal Denaturation ◽  
High Affinity ◽  
Naturally Occurring ◽  
Triostin A ◽  
Bifunctional Compound

The synthesis of a new bifunctional compound in which two aminoacridine chromophores are linked by the bicyclic depsipeptidic backbone of des-N-tetramethylTriostin A is described. The molecule, bis-[(9-acridinyl)-D-seryl-L-alanyl-L-cysteinyl-L-valine] dilactone disulphide, structurally analogous to the antibiotic anti-tumour drug Triostin A, is shown to possess a high affinity to DNA and to act as a bis-intercalator on the basis of spectroscopic, viscosimetric and thermal-denaturation studies. This model constitutes the first attempt of a synergic association between a peptidic moiety that mimics a naturally occurring drug and aminoacridine, the two parts themselves each exhibiting a high affinity for the DNA target.

scholarly journals Bifunctional intercalation and sequence specificity in the binding of quinomycin and triostin antibiotics to deoxyribonucleic acid

1978 ◽  
Vol 173 (1) ◽  
pp. 115-128 ◽  
Author(s):  
J S Lee ◽  
M J Waring
Keyword(s):  
Binding Constant ◽  
Binding Constants ◽  
Sequence Specificity ◽  
Peptide Antibiotics ◽  
Naturally Occurring ◽  
Binding Isotherms ◽  
Triostin A ◽  
Specificity Pattern ◽  
Intercalating Agents ◽  
Binding Curves

Quinomycin C, triostin A and triostin C are peptide antibiotics of the quinoxaline family, of which echinomycin (quinomycin A) is also a member. They all remove and reverse the supercoiling of closed circular duplex DNA from bacteriophage PM2 in the fashion characteristic of intercalating drugs, and the unwinding angle at I 0.01 is, in all cases, almost twice that of ethidium. Thus, as with echinomycin, they can be characterized as bifunctional intercalating agents. For the triostins this conclusion has been confirmed by measurements of changes in the viscosity of sonicated rod-like DNA fragments; the helix extension was found to be almost double that expected for a simple monofunctional intercalation process. For triostin A, further evidence for bifunctionality was derived from the cross-over point of binding isotherms to nicked circular and closed circular bacteriophage-PM2DNA. Binding curves for the interaction of quinomycin C and triostin A with a variety of synthetic and naturally occurring nucleic acids were determined by solvent-partition analysis, but triostin C was too insoluble in aqueous solution to make this method applicable. For quinomycin C the highest binding constant was found with Micrococcus lysodeikticus DNA, and its pattern of specificity among natural DNA species was broadly similar to that of echinomycin, although the binding constants were 2–6 times as large. For triostin A the highest binding constant was again found for M. lysodeikticus DNA, but the specificity pattern was quite different from that of the quinomycins. In particular, triostin A bound better to poly(dA-dT) than to the poly(dG-dC) whereas this order was reversed for quinomycin C. There was also evidence that the binding to poly(dA-dT) might be co-operative in nature. No significant interaction could be detected with poly(dA).poly(dT) or with RNA from Escherichia coli. Poly(dG).poly(dC) gave variable results, depending on the source of the polymer. The different patterns of specificity displayed by the quinomycins and triostins are tentatively ascribed to differences in their conformations in solution.

Thermal Denaturation:  A Method to Rank Slow Binding, High-Affinity P38α MAP Kinase Inhibitors

2003 ◽  
Vol 46 (22) ◽  
pp. 4669-4675 ◽  
Author(s):  
Rachel R. Kroe ◽  
John Regan ◽  
Al Proto ◽  
Gregory W. Peet ◽  
Tapon Roy ◽  
...  
Keyword(s):  
Map Kinase ◽  
Thermal Denaturation ◽  
Kinase Inhibitors ◽  
High Affinity ◽  
P38α Map Kinase

scholarly journals The accumulation and compartmentalization of isometamidium chloride in Trypanosoma congolense, monitored by its intrinsic fluorescence

1995 ◽  
Vol 312 (1) ◽  
pp. 319-327 ◽  
Author(s):  
J M Wilkes ◽  
A S Peregrine ◽  
D Zilberstein
Keyword(s):  
Binding Sites ◽  
Thermal Denaturation ◽  
Specific Binding ◽  
Trypanosoma Congolense ◽  
Emission Peak ◽  
Intrinsic Fluorescence ◽  
Drug Uptake ◽  
Membrane Transporter ◽  
High Affinity ◽  
Dnase 1

Interaction of the trypanocide isometamidium chloride with components of Trypanosoma congolense results in characteristic shifts in the intrinsic fluorescence of the drug. The specificity of this interaction was investigated by analysing the effects of various physicochemical manipulations on its fluorescence properties. The characteristic shifts involved a preferential increase in the intensity of one emission peak over the other, resulting in a systematic increase in the ratio of fluorescence intensities. These effects were apparently due to constraints on fluorophore free rotation in the solution (that is, viscosity). Purified DNA produced similar effects in a saturable manner displaying high affinity for the drug, indicating that the constraint involves binding of the drug to high-affinity binding sites within the DNA. Such binding sites were demonstrated in lysates derived from trypanosomal cells. The binding sites were associated with macromolecular species (M(r) > 12000), and were partly disrupted by thermal denaturation and proteolysis. Treatment with DNase 1 produced high levels of disruption of the binding sites (> 85%), indicating an involvement of DNA in the binding. BSA demonstrated weak non-specific binding of the drug. Entry of drug into live trypanosomal cells (monitored by 14C-labelled drug uptake) was paralleled by fluorescence shifts observed under comparable conditions of drug concentration and buffer conditions. Both systems (fluorescence shifts and accumulation of labelled drug) indicated the presence of a saturable membrane transporter with high affinity for the drug. We conclude that monitoring the fluorescence shifts of isometamidium constitutes a sensitive and highly specific probe for entry of the drug into trypanosomal cells, thereby enabling resolution of the transport events involved.

scholarly journals Nanobody Repertoires for Exposing Vulnerabilities of SARS-CoV-2

2021 ◽  
Author(s):  
Fred D Mast ◽  
Peter C Fridy ◽  
Natalia E Ketaren ◽  
Junjie Wang ◽  
Erica Y Jacobs ◽  
...  
Keyword(s):  
Airway Epithelial Cells ◽  
Spike Protein ◽  
Great Promise ◽  
Antigen Binding ◽  
Receptor Binding Domain ◽  
Airway Epithelial ◽  
Viral Neutralization ◽  
High Affinity ◽  
Naturally Occurring ◽  
Large Repertoire

Despite the great promise of vaccines, the COVID-19 pandemic is ongoing and future serious outbreaks are highly likely, so that multi-pronged containment strategies will be required for many years. Nanobodies are the smallest naturally occurring single domain antigen binding proteins identified to date, possessing numerous properties advantageous to their production and use. We present a large repertoire of high affinity nanobodies against SARS-CoV-2 Spike protein with excellent kinetic and viral neutralization properties, which can be strongly enhanced with oligomerization. This repertoire samples the epitope landscape of the Spike ectodomain inside and outside the receptor binding domain, recognizing a multitude of distinct epitopes and revealing multiple neutralization targets of pseudoviruses and authentic SARS-CoV-2, including in primary human airway epithelial cells. Combinatorial nanobody mixtures show highly synergistic activities, and are resistant to mutational escape and emerging viral variants of concern. These nanobodies establish an exceptional resource for superior COVID-19 prophylactics and therapeutics.

scholarly journals Targeting naturally occurring epitope variants of hepatitis C virus with high-affinity T-cell receptors

2017 ◽  
Vol 98 (3) ◽  
pp. 374-384 ◽  
Author(s):  
Huajun Zhang ◽  
Jianbing Zhang ◽  
Lei Chen ◽  
Zhiming Weng ◽  
Ye Tian ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
T Cell ◽  
T Cell Receptors ◽  
Cell Receptors ◽  
High Affinity ◽  
Naturally Occurring

scholarly journals Engineered Trimeric ACE2 Binds and Locks “Three-up” Spike Protein to Potently Inhibit SARS-CoVs and Mutants

2020 ◽  
Author(s):  
Liang Guo ◽  
Wenwen Bi ◽  
Xinling Wang ◽  
Wei Xu ◽  
Renhong Yan ◽  
...  
Keyword(s):  
De Novo ◽  
Spike Protein ◽  
Protein Scaffold ◽  
High Affinity ◽  
Naturally Occurring ◽  
Native Sequence

AbstractSARS-CoV-2 enters cells via ACE-2, which binds the spike protein with moderate affinity. Despite a constant background mutational rate, the virus must retain binding with ACE2 for infectivity, providing a conserved constraint for SARS-CoV-2 inhibitors. To prevent mutational escape of SARS-CoV-2 and to prepare for future related coronavirus outbreaks, we engineered a de novo trimeric ACE2 (T-ACE2) protein scaffold that binds the trimeric spike protein with extremely high affinity (KD < 1 pM), while retaining ACE2 native sequence. T-ACE2 potently inhibits all tested pseudotyped viruses including SARS-CoV-2, SARS-CoV, eight naturally occurring SARS-CoV-2 mutants, two SARSr-CoVs as well as authentic SARS-CoV-2. The cryo-EM structure reveals that T-ACE2 can induce the transit of spike protein to “three-up” RBD conformation upon binding. T-ACE2 thus represents a promising class of broadly neutralizing proteins against SARS-CoVs and mutants.

Physicochemical Characteristics of a Thermostable Gellan Lyase from Geobacillus stearothermophilus 98

2010 ◽  
Vol 65 (3-4) ◽  
pp. 231-238 ◽  
Author(s):  
Anna Derekova ◽  
Miroslava Atanassova ◽  
Petya Christova ◽  
Bojidar Tchorbanov ◽  
Alexandra Shosheva ◽  
...  
Keyword(s):  
Thermal Denaturation ◽  
Kinetic Studies ◽  
Physicochemical Characteristics ◽  
Thermophilic Bacterium ◽  
Geobacillus Stearothermophilus ◽  
Scanning Calorimetry ◽  
Single Product ◽  
Capillary Gel Electrophoresis ◽  
High Affinity ◽  
Cooperative Transition

A purified thermostable gellan lyase, produced by a thermophilic bacterium, Geobacillus stearothermophilus 98, was characterized in relation to its physicochemical properties. The gellan lyase was established to have a molecular weight of 216 kDa, defined by capillary gel electrophoresis. Amino acid analysis revealed high quantities of Lys, His, Ala, Val, Ile, Glx, and Pro residues. The circular dichroism revealed 45% β-structure and practically lack of α-spiral domains. Kinetic studies showed high affinity of the enzyme to gellan as a substrate (Km = 0.21 μM). The thermal denaturation investigated by cicular dichroism showed a highly cooperative transition with a midpoint (Tm) at about 75 °C. A single product was identified after enzyme action on gellan. Large exothermic aggregation near Tm was observed by differential scanning calorimetry. Two types of gellan lyase crystals were reproducibly isolated.

scholarly journals Fluorescence and circular-dichroism studies on the Streptomyces R61 dd-carboxypeptidase–transpeptidase. Penicillin binding by the enzyme

1973 ◽  
Vol 135 (3) ◽  
pp. 493-505 ◽  
Author(s):  
Manuel Nieto ◽  
Harold R. Perkins ◽  
Jean-Marie Frère ◽  
Jean-Marie Ghuysen
Keyword(s):  
Circular Dichroism ◽  
Thermal Denaturation ◽  
Penicillin G ◽  
Affinity Binding ◽  
Guanidinium Chloride ◽  
Positive Maximum ◽  
Peptide Substrates ◽  
High Affinity ◽  
Circular Dichroïsm ◽  
New Hypothesis

The circular dichroism of the dd-carboxypeptidase–transpeptidase from Streptomyces R61 shows in the near u.v. a set of weak extrema at 289nm (positive) and at 282, 275 and 268nm (all negative). In the far u.v. it shows negative extrema at 217–218 and 208nm, crossover at 202nm and a positive maximum at about 194nm. The u.v. absorption of the enzyme shows it to contain tyrosine and tryptophan in approx. 3.4:1 ratio. The enzyme is fluorescent with a maximum emission at 318–320nm. The near-u.v. circular dichroism of the protein is extensively affected by binding of penicillin G, but the far u.v. is unaffected. Binding of the antibiotic also causes quenching of the fluorescence of the enzyme. The latter effect has been used to study the binding of penicillin G to the enzyme and the influence exerted upon it by salts, denaturants and peptide substrates and inhibitors. High-affinity binding of penicillin appears to be comparatively slow and reversible, and can occur under conditions in which the protein is enzymically inactive. The thermal denaturation of the enzyme in guanidinium chloride at pH7 is affected by binding of the antibiotic. The presence of even large concentrations of β-mercaptoethanol neither impaired the activity of the enzyme nor prevented its inhibition by penicillin G or cephalosporin C. A new hypothesis for the molecular mechanism of the interaction of the enzyme with penicillin is proposed.

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