scholarly journals A pH-dependent bolt involving cytosine bases located in the lateral loops of antiparallel G-quadruplex structures within the SMARCA4 gene promotor

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Sanae Benabou ◽  
Stefania Mazzini ◽  
Anna Aviñó ◽  
Ramon Eritja ◽  
Raimundo Gargallo
Keyword(s):  
Base Pair ◽  
Thermodynamic Description ◽  
Size Exclusion ◽  
Spectroscopic Techniques ◽  
G Quadruplex ◽  
Exclusion Chromatography ◽  
Additional Stability ◽  
Gene Promotor ◽  
Ph Dependent ◽  
Melting Experiments

Abstract Some lung and ovarian tumors are connected to the loss of expression of SMARCA4 gene. In its promoter region, a 44-nucleotides long guanine sequence prone to form G-quadruplex structures has been studied by means of spectroscopic techniques (circular dichroism, molecular absorption and nuclear magnetic resonance), size exclusion chromatography and multivariate analysis. The results have shown that the central 21-nucleotides long sequence comprising four guanine tracts of disparate length is able to fold into a pH-dependent ensemble of G-quadruplex structures. Based on acid-base titrations and melting experiments of wild and mutated sequences, the formation of a C·C+ base pair between cytosine bases present at the two lateral loops is shown to promote a reduction in conformational heterogeneity, as well as an increase in thermal stability. The formation of this base pair is characterized by a pKa value of 7.1 ± 0.2 at 20 °C and 150 mM KCl. This value, higher than those usually found in i-motif structures, is related to the additional stability provided by guanine tetrads in the G-quadruplex. To our knowledge, this is the first thermodynamic description of this base pair in loops of antiparallel G-quadruplex structures.

scholarly journals Structural and Functional Aspects of G-Quadruplex Aptamers Which Bind a Broad Range of Influenza A Viruses

Biomolecules ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 119
Author(s):  
Anastasia A. Novoseltseva ◽  
Nikita M. Ivanov ◽  
Roman A. Novikov ◽  
Yaroslav V. Tkachev ◽  
Dmitry A. Bunin ◽  
...  
Keyword(s):  
Influenza A ◽  
Specific Binding ◽  
Dna Aptamer ◽  
Size Exclusion ◽  
Influenza A Viruses ◽  
Strain Specificity ◽  
Influenza Hemagglutinin ◽  
Functional Aspects ◽  
G Quadruplex ◽  
Exclusion Chromatography

An aptamer is a synthetic oligonucleotide with a unique spatial structure that provides specific binding to a target. To date, several aptamers to hemagglutinin of the influenza A virus have been described, which vary in affinity and strain specificity. Among them, the DNA aptamer RHA0385 is able to recognize influenza hemagglutinins with highly variable sequences. In this paper, the structure of RHA0385 was studied by circular dichroism spectroscopy, nuclear magnetic resonance, and size-exclusion chromatography, demonstrating the formation of a parallel G-quadruplex structure. Three derivatives of RHA0385 were designed in order to determine the contribution of the major loop to affinity. Shortening of the major loop from seven to three nucleotides led to stabilization of the scaffold. The affinities of the derivatives were studied by surface plasmon resonance and an enzyme-linked aptamer assay on recombinant hemagglutinins and viral particles, respectively. The alterations in the loop affected the binding to influenza hemagglutinin, but did not abolish it. Contrary to aptamer RHA0385, two of the designed aptamers were shown to be conformationally homogeneous, retaining high affinities and broad binding abilities for both recombinant hemagglutinins and whole influenza A viruses.

scholarly journals New G-Quadruplex-Forming Oligodeoxynucleotides Incorporating a Bifunctional Double-Ended Linker (DEL): Effects of DEL Size and ODNs Orientation on the Topology, Stability, and Molecularity of DEL-G-Quadruplexes

Molecules ◽  
2019 ◽  
Vol 24 (3) ◽  
pp. 654 ◽  
Author(s):  
Maria Marzano ◽  
Andrea Falanga ◽  
Stefano D’Errico ◽  
Brunella Pinto ◽  
Giovanni Roviello ◽  
...  
Keyword(s):  
Size Exclusion Chromatography ◽  
Structural Characterization ◽  
Size Exclusion ◽  
Biological Processes ◽  
Dna Nanostructures ◽  
Nmr Studies ◽  
Molecular Weights ◽  
G Quadruplex ◽  
Exclusion Chromatography

G-quadruplexes (G4s) are unusual secondary structures of DNA occurring in guanosine-rich oligodeoxynucleotide (ODN) strands that are extensively studied for their relevance to the biological processes in which they are involved. In this study, we report the synthesis of a new kind of G4-forming molecule named double-ended-linker ODN (DEL-ODN), in which two TG4T strands are attached to the two ends of symmetric, non-nucleotide linkers. Four DEL-ODNs differing for the incorporation of either a short or long linker and the directionality of the TG4T strands were synthesized, and their ability to form G4 structures and/or multimeric species was investigated by PAGE, HPLC–size-exclusion chromatography (HPLC–SEC), circular dichroism (CD), and NMR studies in comparison with the previously reported monomeric tetra-ended-linker (TEL) analogues and with the corresponding tetramolecular species (TG4T)4. The structural characterization of DEL-ODNs confirmed the formation of stable, bimolecular DEL-G4s for all DEL-ODNs, as well as of additional DEL-G4 multimers with higher molecular weights, thus suggesting a way towards the obtainment of thermally stable DNA nanostructures based on reticulated DEL-G4s.

scholarly journals Variable-Temperature Size Exclusion Chromatography for the Study of the Structural Changes in G-Quadruplex

2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Sanae Benabou ◽  
Ramon Eritja ◽  
Raimundo Gargallo
Keyword(s):  
Size Exclusion Chromatography ◽  
Promoter Region ◽  
Structural Changes ◽  
Variable Temperature ◽  
Size Exclusion ◽  
G Quadruplex ◽  
Conformational Equilibria ◽  
Exclusion Chromatography ◽  
In The Wild ◽  
Circular Dichroïsm

The conformational equilibria of a guanine-rich sequence found at the promoter region of the human c-kit oncogene are studied by means of circular dichroism spectroscopy (CD) and variable-temperature size exclusion chromatography (SEC). It is shown that the wild sequence ckit21 exists as a mixture of monomeric and multimeric G-quadruplexes. Appropriate mutation of several bases in the wild sequence produces the shift from parallel to antiparallel G-quadruplex, as well as the disappearance of multimeric species. The shift from the antiparallel to the parallel conformation induced by temperature is reflected in both CD and SEC profiles.

scholarly journals Organic Foulants Characteristics in Membrane Bioreactor

2017 ◽  
Vol 5 (1) ◽  
Author(s):  
R. K. Aryal ◽  
S. Vigneswaran ◽  
J. Lebegue ◽  
H. K. Shon ◽  
J. Kandasamy ◽  
...  
Keyword(s):  
Membrane Bioreactor ◽  
Membrane Surface ◽  
Aeration Rate ◽  
Size Exclusion ◽  
Spectroscopic Techniques ◽  
Exclusion Chromatography ◽  
Naoh Solution ◽  
Bioreactor System ◽  
Microbial Products ◽  
Flat Sheet Membrane

A laboratory scale side stream membrane bioreactor system with flat sheet membrane was operated for 5–days run at three different aeration rates (100, 200 and 300 L/h). The organic foulants deposited on the membrane surface was studied after extraction with 5% NaOH solution using three spectroscopic techniques. The IR spectra showed no distinct similarity in peaks among the three. The fluorescence spectra showed increase of soluble microbial products in foulant with decrease of aeration rate. This was supported by the size exclusion chromatography in which biopolymers concentration in fouling decreased with increasing aeration rate.

scholarly journals Stability-Indicating Analytical Approach for Stability Evaluation of Lactoferrin

Pharmaceutics ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1065
Author(s):  
Nika Osel ◽  
Timeja Planinšek Parfant ◽  
Albin Kristl ◽  
Robert Roškar
Keyword(s):  
Analytical Approach ◽  
Degradation Products ◽  
Reversed Phase ◽  
Size Exclusion ◽  
Spectroscopic Techniques ◽  
Stability Evaluation ◽  
Stability Indicating ◽  
Exclusion Chromatography ◽  
Quality Of Products ◽  
The Stability

Lactoferrin is a multifunctional iron-binding glycoprotein in milk. Due to its potential for the treatment of various diseases, interest in products containing lactoferrin is increasing. However, as a protein, it is prone to degradation, which critically affects the quality of products. Therefore, the main purpose of our work was to develop a stability-indicating analytical approach for stability evaluation of lactoferrin. We were focused on two complementary methods: reversed-phase and size-exclusion chromatography. The stability-indicating nature of the selected methods was confirmed. They were successfully validated by following the ICH guidelines and applied to preliminary lactoferrin stability studies. Up to three degradation products, as well as aggregates and fragments of lactoferrin, were detected in various samples using complementary reversed-phase and size-exclusion chromatographic methods. The analytical approach was additionally extended with three spectroscopic techniques (absorbance, intrinsic fluorescence, and bicinchoninic acid method), which may provide valuable complementary information in some cases. The presented analytical approach allows the stability evaluation of lactoferrin in various samples, including the ability to detect differences in its degradation mechanisms. Furthermore, it has the potential to be used for the quality control of products containing lactoferrin.

scholarly journals Insights into the G-rich VEGF-binding aptamer V7t1: when two G-quadruplexes are better than one!

2019 ◽  
Vol 47 (15) ◽  
pp. 8318-8331 ◽  
Author(s):  
Federica Moccia ◽  
Claudia Riccardi ◽  
Domenica Musumeci ◽  
Serena Leone ◽  
Rosario Oliva ◽  
...  
Keyword(s):  
Size Exclusion ◽  
Biophysical Characterization ◽  
Monomeric Species ◽  
Conformational Preferences ◽  
G Quadruplex ◽  
Exclusion Chromatography ◽  
Extracellular Environment ◽  
Annealed Oligonucleotide ◽  
Better Than

Abstract The G-quadruplex-forming VEGF-binding aptamer V7t1 was previously found to be highly polymorphic in a K+-containing solution and, to restrict its conformational preferences to a unique, well-defined form, modified nucleotides (LNA and/or UNA) were inserted in its sequence. We here report an in-depth biophysical characterization of V7t1 in a Na+-rich medium, mimicking the extracellular environment in which VEGF targeting should occur, carried out combining several techniques to analyse the conformational behaviour of the aptamer and its binding to the protein. Our results demonstrate that, in the presence of high Na+ concentrations, V7t1 behaves in a very different way if subjected or not to annealing procedures, as evidenced by native gel electrophoresis, size exclusion chromatography and dynamic light scattering analysis. Indeed, not-annealed V7t1 forms both monomeric and dimeric G-quadruplexes, while the annealed oligonucleotide is a monomeric species. Remarkably, only the dimeric aptamer efficiently binds VEGF, showing higher affinity for the protein compared to the monomeric species. These findings provide new precious information for the development of improved V7t1 analogues, allowing more efficient binding to the cancer-related protein and the design of effective biosensors or theranostic devices based on VEGF targeting.

CHARACTERIZATION OF A NEW LCAT MUTATION CAUSING FAMILIAL LCAT DEFICIENCY (FLD) AND THE ROLE OF APOE AS A MODIFIER GENE OF THE FLD PHENOTYPE

2009 ◽  
Vol 32 (6S) ◽  
pp. 3
Author(s):  
A Baass ◽  
H Wassef ◽  
M Tremblay ◽  
L Bernier ◽  
R Dufour ◽  
...  
Keyword(s):  
Modifier Gene ◽  
Size Exclusion ◽  
Plasma Lipoproteins ◽  
Lipoprotein Profile ◽  
Exclusion Chromatography ◽  
Low Hdl ◽  
Lcat Deficiency ◽  
Apoe Genotypes

Introduction: LCAT (lecithin:cholesterol acyltransferase ) is an enzyme which plays an essential role in cholesterol esterification and reverse cholesterol transport. Familial LCAT deficiency (FLD) is a disease characterized by a defect in LCAT resulting in extremely low HDL-C, premature corneal opacities, anemia as well as proteinuria and renal failure. Method: We have identified two brothers presenting characteristics of familial LCAT deficiency. We sequenced the LCAT gene, measured the lipid profile as well as the LCAT activity in 15 members of this kindred. We also characterized the plasma lipoproteins by agarose gel electrophoresis and size exclusion chromatography and sequenced several candidate genes related to dysbetalipoproteinemia in this family. Results: We have identified the first French Canadian kindred with familial LCAT deficiency. Two brothers affected by FLD, were homozygous for a novel LCAT mutation. This c.102delG mutation occurs at the codon for His35 causing a frameshift that stops transcription at codon 61 abolishing LCAT enzymatic activity both in vivo and in vitro. It has a dramatic effect on the lipoprotein profile, with an important reduction of HDL-C in both heterozygotes (22%) and homozygotes (88%) and a significant decrease in LDL-C in heterozygotes (35%) as well as homozygotes (58%). Furthermore, the lipoprotein profile differed markedly between the two affected brothers who had different APOE genotypes. We propose that APOE could be an important modifier gene explaining heterogeneity in lipoprotein profiles observed among FLD patients. Our results suggest that a LCAT-/- genotype associated with an APOE ?2 allele could be a novel mechanism leading to dysbetalipoproteinemia.

Structure of a shear-thickening polysaccharide extracted from the New Zealand black tree fern, Cyathea medullaris

2020 ◽  
Author(s):  
M Wee ◽  
M Mastrangelo ◽  
Susan Carnachan ◽  
Ian Sims ◽  
K Goh
Keyword(s):  
New Zealand ◽  
Uronic Acid ◽  
Average Molecular Weight ◽  
Shear Thickening ◽  
Water Soluble ◽  
Size Exclusion ◽  
Resonance Spectroscopy ◽  
Tree Fern ◽  
13C Nuclear Magnetic Resonance ◽  
Exclusion Chromatography

A shear-thickening water-soluble polysaccharide was purified from mucilage extracted from the fronds of the New Zealand black tree fern (Cyathea medullaris or 'mamaku' in Māori) and its structure characterised. Constituent sugar analysis by three complementary methods, combined with linkage analysis (of carboxyl reduced samples) and 1H and 13C nuclear magnetic resonance spectroscopy (NMR) revealed a glucuronomannan comprising a backbone of 4-linked methylesterified glucopyranosyl uronic acid and 2-linked mannopyranosyl residues, branched at O-3 of 45% and at both O-3 and O-4 of 53% of the mannopyranosyl residues with side chains likely comprising terminal xylopyranosyl, terminal galactopyranosyl, non-methylesterified terminal glucopyranosyl uronic acid and 3-linked glucopyranosyl uronic acid residues. The weight-average molecular weight of the purified polysaccharide was ~1.9×106Da as determined by size-exclusion chromatography coupled with multi-angle laser light scattering (SEC-MALLS). The distinctive rheological properties of this polysaccharide are discussed in relation to its structure. © 2014 Elsevier B.V.

Sign in / Sign up

Export Citation Format

Share Document