scholarly journals Structure-activity analysis of truncated albumin-binding domains suggests new lead constructs for potential therapeutic delivery

2020 ◽  
Vol 295 (34) ◽  
pp. 12143-12152
Author(s):  
Conan K. Wang ◽  
Anna S. Amiss ◽  
Joachim Weidmann ◽  
David J. Craik
Keyword(s):  
Structural Integrity ◽  
Cholesterol Metabolism ◽  
Activity Analysis ◽  
Titration Calorimetry ◽  
Albumin Binding ◽  
Binding Studies ◽  
Binding Domains ◽  
Structure Activity ◽  
Wide Range

Rapid clearance by renal filtration is a major impediment to the translation of small bioactive biologics into drugs. To extend serum t1/2, a commonly used approach is to attach drug leads to the G-related albumin-binding domain (ABD) to bind albumin and evade clearance. Despite the success of this approach in extending half-lives of a wide range of biologics, it is unclear whether the existing constructs are optimized for binding and size; any improvements along these lines could lead to improved drugs. Characterization of the biophysics of binding of an ABD to albumin in solution could shed light on this question. Here, we examine the binding of an ABD to human serum albumin using isothermal titration calorimetry and assess the structural integrity of the ABD using CD, NMR, and molecular dynamics. A structure-activity analysis of truncations of the ABD suggests that downsized variants could replace the full-length domain. Reducing size could have the benefit of reducing potential immunogenicity problems. We further showed that one of these variants could be used to design a bifunctional molecule with affinity for albumin and a serum protein involved in cholesterol metabolism, PCSK9, demonstrating the potential utility of these fragments in the design of cholesterol-lowering drugs. Future work could extend these in vitro binding studies to other ABD variants to develop therapeutics. Our study presents new understanding of the solution structural and binding properties of ABDs, which has implications for the design of next-generation long-lasting therapeutics.

scholarly journals Two peptides targeting endothelial receptors are internalized into murine brain endothelial cells

PLoS ONE ◽  
2021 ◽  
Vol 16 (4) ◽  
pp. e0249686
Author(s):  
Diána Hudecz ◽  
Sara Björk Sigurdardóttir ◽  
Sarah Christine Christensen ◽  
Casper Hempel ◽  
Andrew J. Urquhart ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Transferrin Receptor ◽  
Vesicular Transport ◽  
Brain Diseases ◽  
Brain Endothelial Cells ◽  
Binding Studies ◽  
Wide Range ◽  
Murine Brain ◽  
The Brain

The blood-brain barrier (BBB) is one of the main obstacles for therapies targeting brain diseases. Most macromolecules fail to pass the tight BBB, formed by brain endothelial cells interlinked by tight junctions. A wide range of small, lipid-soluble molecules can enter the brain parenchyma via diffusion, whereas macromolecules have to transcytose via vesicular transport. Vesicular transport can thus be utilized as a strategy to deliver brain therapies. By conjugating BBB targeting antibodies and peptides to therapeutic molecules or nanoparticles, it is possible to increase uptake into the brain. Previously, the synthetic peptide GYR and a peptide derived from melanotransferrin (MTfp) have been suggested as candidates for mediating transcytosis in brain endothelial cells (BECs). Here we study uptake, intracellular trafficking, and translocation of these two peptides in BECs. The peptides were synthesized, and binding studies to purified endocytic receptors were performed using surface plasmon resonance. Furthermore, the peptides were conjugated to a fluorophore allowing for live-cell imaging studies of their uptake into murine brain endothelial cells. Both peptides bound to low-density lipoprotein receptor-related protein 1 (LRP-1) and the human transferrin receptor, while lower affinity was observed against the murine transferrin receptor. The MTfp showed a higher binding affinity to all receptors when compared to the GYR peptide. The peptides were internalized by the bEnd.3 mouse endothelial cells within 30 min of incubation and frequently co-localized with endo-lysosomal vesicles. Moreover, our in vitro Transwell translocation experiments confirmed that GYR was able to cross the murine barrier and indicated the successful translocation of MTfp. Thus, despite binding to endocytic receptors with different affinities, both peptides are able to transcytose across the murine BECs.

scholarly journals Novel Azolylalkyloxy Compounds with Antipicornaviral Activity

1995 ◽  
Vol 6 (4) ◽  
pp. 245-254 ◽  
Author(s):  
M. D. Abel ◽  
A. D. Cameron ◽  
C. M. Ha ◽  
C. A. Koski ◽  
H. T. Luu ◽  
...  
Keyword(s):  
Vitro Activity ◽  
In Vitro Activity ◽  
Optimal Length ◽  
Structure Activity Relationships ◽  
Structure Activity ◽  
Wide Range ◽  
Highly Effective ◽  
Qualitative Structure

A novel series of azolylalkyloxy compounds was designed, synthesized and evaluated for antipicornaviral activity. Several of the compounds exhibited in vitro activity comparable to that of Disoxarll. An investigation of qualitative structure-activity relationships indicated that the optimal length of the alkyI chain is six or seven carbon atoms, with seven being marginally superior. The effect of different azole moieties on activity was relatively small, with 3-methylisoxazole and 4-methylthiazole being most effective. The nature of the oxy substituent was found to be extremely important for antipicornaviral activity. The 2-dibenzofuryl group proved to be the most effective oxy substituent for this class of compounds. Compounds 11 and 22, combining dibenzofuran with 3-methylisoxazole and 4-methylthiazole, respectively, were highly effective in vitro against a wide range of human rhinoviruses as well as several enteroviruses.

scholarly journals An Interpenetrating Alginate/Gelatin Network for Three-Dimensional (3D) Cell Cultures and Organ Bioprinting

Molecules ◽  
2020 ◽  
Vol 25 (3) ◽  
pp. 756 ◽  
Author(s):  
Qiuhong Chen ◽  
Xiaohong Tian ◽  
Jun Fan ◽  
Hao Tong ◽  
Qiang Ao ◽  
...  
Keyword(s):  
Cell Cultures ◽  
Structural Integrity ◽  
Polymer Network ◽  
Three Dimensional ◽  
Biochemical Properties ◽  
Interpenetrating Polymer Network ◽  
3D Cell Cultures ◽  
Wide Range ◽  
The Individual

Crosslinking is an effective way to improve the physiochemical and biochemical properties of hydrogels. In this study, we describe an interpenetrating polymer network (IPN) of alginate/gelatin hydrogels (i.e., A-G-IPN) in which cells can be encapsulated for in vitro three-dimensional (3D) cultures and organ bioprinting. A double crosslinking model, i.e., using Ca2+ to crosslink alginate molecules and transglutaminase (TG) to crosslink gelatin molecules, is exploited to improve the physiochemical, such as water holding capacity, hardness and structural integrity, and biochemical properties, such as cytocompatibility, of the alginate/gelatin hydrogels. For the sake of convenience, the individual ionic (i.e., only treatment with Ca2+) or enzymatic (i.e., only treatment with TG) crosslinked alginate/gelatin hydrogels are referred as alginate-semi-IPN (i.e., A-semi-IPN) or gelatin-semi-IPN (i.e., G-semi-IPN), respectively. Tunable physiochemical and biochemical properties of the hydrogels have been obtained by changing the crosslinking sequences and polymer concentrations. Cytocompatibilities of the obtained hydrogels are evaluated through in vitro 3D cell cultures and bioprinting. The double crosslinked A-G-IPN hydrogel is a promising candidate for a wide range of biomedical applications, including bioartificial organ manufacturing, high-throughput drug screening, and pathological mechanism analyses.

Xenin relaxes precontracted rat ileal smooth muscle in vitro: Structure activity relationship and binding studies

1995 ◽  
Vol 108 (4) ◽  
pp. A585
Author(s):  
A. Clemens ◽  
S. Katsoulis ◽  
R. Nustede ◽  
K. Seyfarth ◽  
C. Morys-Wortman ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Structure Activity Relationship ◽  
Activity Relationship ◽  
Binding Studies ◽  
Structure Activity

Relaxant effect of xenin on rat ileum is mediated by apamin-sensitive neurotensin-type receptors

1997 ◽  
Vol 272 (1) ◽  
pp. G190-G196 ◽  
Author(s):  
A. Clemens ◽  
S. Katsoulis ◽  
R. Nustede ◽  
J. Seebeck ◽  
K. Seyfarth ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Channel Blocker ◽  
Muscle Cells ◽  
K Channel ◽  
Binding Studies ◽  
Relaxant Effect ◽  
Structure Activity ◽  
Neuromedin N

The action of xenin, a novel 25-residue peptide of the neurotensin (NT)/xenopsin family, was investigated in isolated rat ileal muscle strips and in dispersed longitudinal smooth muscle cells of rat small intestine in vitro. Xenin relaxes KCl-precontracted ileal strips dose dependently (1 nM-3 microM). The order of potency of the investigated peptides was as follows: xenopsin = NT = xenin > neuromedin N. Kinetensin was inactive. Tetrodotoxin, hexamethonium, tetraethylammonium, 4-aminopyridine, and NG-nitro-L-arginine did not influence the relaxant effects of xenin or NT, whereas the K+ channel blocker apamin nearly abolished their effects. Desensitization against one of the peptides or blockade of NT receptors by SR-48692 prevented the effect of xenin and NT. Structure-activity experiments revealed that the COOH-terminal part of the molecules of xenin and NT is essential for biological activity. Experiments with isolated dispersed smooth muscle cells and binding studies on intestinal smooth muscle cell membranes confirmed and extended the results obtained with muscle strips. In conclusion, xenin relaxes rat ileal smooth muscle via a muscular NT-type apamin-sensitive receptor.

α2-Macroglobulin, the main serum antiprotease, binds β2-microglobulin, the light chain of the class I major histocompatibility complex, which is involved in human disease

2000 ◽  
Vol 98 (4) ◽  
pp. 427-433 ◽  
Author(s):  
Annie GOUIN-CHARNET ◽  
Daniel LAUNE ◽  
Claude GRANIER ◽  
Jean-Claude MANI ◽  
Bernard PAU ◽  
...  
Keyword(s):  
Major Histocompatibility Complex ◽  
Clinical Findings ◽  
Class I ◽  
Major Histocompatibility ◽  
Binding Studies ◽  
Β2 Microglobulin ◽  
Histocompatibility Complex ◽  
Α2 Macroglobulin ◽  
Wide Range

β2-Microglobulin, a 12 kDa protein forming part of the class I HLA (histocompatibility locus antigen) major histocompatibility complex, has been used as a prognosis factor for multiple myeloma and as a marker of renal function, and has been shown to be involved in the pathogenesis of dialysis-related amyloidosis. α2-Macroglobulin has the ability to bind a wide range of physiologically important molecules, thereby influencing their metabolic impact. In this study we show by Western blotting analysis that β2-microglobulin binds to α2-macroglobulin in vitro. This binding was confirmed by BIAcore analysis, and was shown by ELISA to be concentration-dependent. The sequences of the binding peptides in the mature β2-microglobulin molecule were identified by Spot multiple peptide synthesis and α2-macroglobulin binding studies. In conclusion, β2-microglobulin interacts specifically with the universal antiprotease a2-macroglobulin. The identification of this interaction brings into question some of the axioms on the metabolism of β2-microglobulin, and may help to explain the clinical findings observed in b2-microglobulin-related diseases.

scholarly journals Antiplasmodial Chalcones Inhibit Sorbitol-Induced Hemolysis of Plasmodium falciparum-Infected Erythrocytes

2004 ◽  
Vol 48 (9) ◽  
pp. 3241-3245 ◽  
Author(s):  
Mei-Lin Go ◽  
Mei Liu ◽  
Prapon Wilairat ◽  
Philip J. Rosenthal ◽  
Kevin J. Saliba ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Erythrocyte Membrane ◽  
Malaria Parasite ◽  
Important Determinant ◽  
Activity Analysis ◽  
Significant Extent ◽  
Active Compounds ◽  
Structure Activity ◽  
Qualitative Structure

ABSTRACT A series of alkoxylated and hydroxylated chalcones previously reported to have antiplasmodial activities in vitro were investigated for their effects on the new permeation pathways induced by the malaria parasite in the host erythrocyte membrane. Of 21 compounds with good antiplasmodial activities (50% inhibitory concentrations [IC50s], ≤20 μM), 8 members were found to inhibit sorbitol-induced lysis of parasitized erythrocytes to a significant extent (≤40% of control values) at a concentration (10 μM) that was close to their antiplasmodial IC50s. Qualitative structure-activity analysis suggested that activity was governed to a greater extent by a substitution on ring B than on ring A of the chalcone template. Most of the active compounds had methoxy or dimethoxy groups on ring B. Considerable variety was permitted on ring A in terms of the electron-donating or -withdrawing property. Lipophilicity did not appear to be an important determinant for activity. Although they are not exceptionally potent as inhibitors (lowest IC50, 1.9 μM), the chalcones compare favorably with other more potent inhibitors in terms of their selective toxicities against plasmodia and their neutral character.

scholarly journals Development of an in vitro tissue culture system for hammer coral (Fimbriaphyllia ancora) ovaries

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yi-Ling Chiu ◽  
Ching-Fong Chang ◽  
Shinya Shikina
Keyword(s):  
Structural Integrity ◽  
Culture Method ◽  
Culture Conditions ◽  
Short Term ◽  
Wide Range ◽  
Fetal Bovine ◽  
Short Term Culture ◽  
Mesenterial Filaments

AbstractIn vitro gonad culture systems have proven useful to investigate intrinsic mechanisms of sexual reproduction in animals. Here we describe development of an in vitro culture method for coral ovaries. Mesenterial tissues containing both ovaries and mesenterial filaments were microscopically isolated from the scleractinian coral, Fimbriaphyllia ancora, and culture conditions were optimized. M199 diluted 10× (10% M199, pH 8.1) and supplemented with 25 mM HEPES and the antibiotics, ampicillin, penicillin and streptomycin, supported oocyte survival and maintained the structural integrity of ovaries during short-term culture (~ 6 days). Addition of a commercial antibiotic–antimycotic solution (Anti–Anti) and fetal bovine serum adversely affected ovary maintenance and caused tissue disintegration. Characterization of cultured ovaries showed that there is no difference in cell proliferation of ovarian somatic cells between culture Days 1 and 6. Moreover, the presence of oogonia and expression of a major yolk protein, vitellogenin, were confirmed in ovaries cultured for 6 days. This system will be useful for studying effects of a wide range of substances on coral oogenesis.

Vitex Diterpenoids: Structural Diversity and Pharmacological Activity

2020 ◽  
Vol 26 (1) ◽  
pp. 138-159 ◽  
Author(s):  
Yanfei Ban ◽  
Tianshuang Xia ◽  
Rui Jing ◽  
Yaoli Guo ◽  
Yiya Geng ◽  
...  
Keyword(s):  
Pharmacological Activity ◽  
Hormone Level ◽  
Biological Activities ◽  
Folk Medicine ◽  
Structural Diversity ◽  
Pharmacological Activities ◽  
Structure Activity ◽  
Wide Range ◽  
Potential Applications

Plants of the genus Vitex (Verbenaceae) are mainly distributed throughout tropical and temperate regions, and many Vitex plants have been traditionally used in folk medicine. Plants of this genus are a rich source of diterpenoids, which not only displayed versatile structural diversity with potential chemotaxonomical significance but also exhibited a wide range of biological activities, mainly including in vitro cytotoxic, antiinflammatory, antimicrobial, hormone level-regulating and antiangiogenic activities. Recently, a series of bioactive diterpenoids, with interesting carbon skeletons, have been reported and gathered considerable interest. This article systematically reviewed diterpenoids isolated from the genus Vitex that appeared in the literature up to December 2018, critically highlighting their structural diversity and pharmacological activities. Up to now, a total of 154 diterpenoids with diverse structures have been isolated and identified from Vitex plants. The authors also summarized the reported structure-activity relationships of those well explored Vitex diterpenoids. Finally, the authors discussed the challenges and potential applications of these diterpenoids in the future.

Solution structure, dynamics and binding studies of a family 11 carbohydrate-binding module from Clostridium thermocellum (CtCBM11)

2013 ◽  
Vol 451 (2) ◽  
pp. 289-300 ◽  
Author(s):  
Aldino Viegas ◽  
João Sardinha ◽  
Filipe Freire ◽  
Daniel F. Duarte ◽  
Ana L. Carvalho ◽  
...  
Keyword(s):  
Clostridium Thermocellum ◽  
Solution Structure ◽  
Close Contact ◽  
Van Der Waals Interactions ◽  
Backbone Dynamics ◽  
Titration Calorimetry ◽  
Carbohydrate Binding ◽  
Binding Studies ◽  
Carbohydrate Binding Modules ◽  
Wide Range

Non-catalytic cellulosomal CBMs (carbohydrate-binding modules) are responsible for increasing the catalytic efficiency of cellulosic enzymes by selectively putting the substrate (a wide range of poly- and oligo-saccharides) and enzyme into close contact. In the present study we carried out an atomistic rationalization of the molecular determinants of ligand specificity for a family 11 CBM from thermophilic Clostridium thermocellum [CtCBM11 (C. thermocellum CBM11)], based on a NMR and molecular modelling approach. We have determined the NMR solution structure of CtCBM11 at 25°C and 50°C and derived information on the residues of the protein that are involved in ligand recognition and on the influence of the length of the saccharide chain on binding. We obtained models of the CtCBM11–cellohexaose and CtCBM11–cellotetraose complexes by docking in accordance with the NMR experimental data. Specific ligand–protein CH-π and Van der Waals interactions were found to be determinant for the stability of the complexes and for defining specificity. Using the order parameters derived from backbone dynamics analysis in the presence and absence of ligand and at 25°C and 50°C, we determined that the protein's backbone conformational entropy is slightly positive. This data in combination with the negative binding entropy calculated from ITC (isothermal titration calorimetry) studies supports a selection mechanism where a rigid protein selects a defined oligosaccharide conformation.

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