scholarly journals Conformation and protein interactions of intramolecular DNA and phosphorothioate four-way junctions

2020 ◽  
pp. 153537022097397
Author(s):  
Maria Troisi ◽  
Mitchell Klein ◽  
Andrew C Smith ◽  
Gaston Moorhead ◽  
Yonatan Kebede ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Therapeutic Potential ◽  
Dnase I ◽  
Protein Recognition ◽  
Exonuclease Iii ◽  
Branched Dna ◽  
Acid Cleavage ◽  
Nuclease Digestion ◽  
Nuclease Resistance ◽  
Exo Iii

The objectives of this study are to evaluate the structure and protein recognition features of branched DNA four-way junctions in an effort to explore the therapeutic potential of these molecules. The classic immobile DNA 4WJ, J1, is used as a matrix to design novel intramolecular junctions including natural and phosphorothioate bonds. Here we have inserted H2-type mini-hairpins into the helical termini of the arms of J1 to generate four novel intramolecular four-way junctions. Hairpins are inserted to reduce end fraying and effectively eliminate potential nuclease binding sites. We compare the structure and protein recognition features of J1 with four intramolecular four-way junctions: i-J1, i-J1(PS1), i-J1(PS2) and i-J1(PS3). Circular dichroism studies suggest that the secondary structure of each intramolecular 4WJ is composed predominantly of B-form helices. Thermal unfolding studies indicate that intramolecular four-way junctions are significantly more stable than J1. The Tm values of the hairpin four-way junctions are 25.2° to 32.2°C higher than the control, J1. With respect to protein recognition, gel shift assays reveal that the DNA-binding proteins HMGBb1 and HMGB1 bind the hairpin four-way junctions with affinity levels similar to control, J1. To evaluate nuclease resistance, four-way junctions are incubated with DNase I, exonuclease III (Exo III) and T5 exonuclease (T5 Exo). The enzymes probe nucleic acid cleavage that occurs non-specifically (DNase I) and in a 5ʹ→3ʹ (T5 Exo) and 3ʹ→5ʹ direction (Exo III). The nuclease digestion assays clearly show that the intramolecular four-way junctions possess significantly higher nuclease resistance than the control, J1.

Synchrony of Digestion of SV40 DNA by Exonuclease III Determined by EM

1975 ◽  
Vol 33 ◽  
pp. 674-675
Author(s):  
Ray Wu ◽  
G. Ruben ◽  
B. Siegel ◽  
P. Spielman ◽  
E. Jay
Keyword(s):  
Dark Field ◽  
Uranyl Acetate ◽  
Enzyme Digestion ◽  
Dna Molecules ◽  
Exonuclease Iii ◽  
Aluminum Films ◽  
Double Stranded Dna ◽  
Before And After ◽  
Exo Iii ◽  
Sv40 Dna

A method for determining long nucleotide sequences of double-stranded DNA is being developed. It involves (a) the synchronous digestion of the DNA from the 3' ends with EL coli exonuclease III (Exo III) followed by (b) resynthesis with labeled nucleotides and DNA polymerase. A crucial factor in the success of this method is the degree to which the enzyme digestion proceeds synchronously under proper conditions of incubation (step a). Dark field EM is used to obtain accurate measurements on the lengths and distribution of the DNA molecules before and after digestion with Exo III, while gel electrophoresis is used in parallel to obtain a mean length for these molecules. It is the measurements on a large enough sample of individual molecules by EM that provides the information on how synchronously the digestion proceeds. For length measurements, the DNA molecules were picked up on 20-30 Å thick carbon-aluminum films, using the aqueous Kleinschmidt technique and stained with 7.5 x 10-5M uranyl acetate in 90% ethanol for 3 minutes.

Ultrasensitive fluorometric biosensor based on Ti3C2 MXenes with Hg2+-triggered exonuclease III-assisted recycling amplification

The Analyst ◽  
2021 ◽  
Author(s):  
Liling Lu ◽  
Xiao Han ◽  
Jingwen Lin ◽  
Yingxin Zhang ◽  
Minghao Qiu ◽  
...  
Keyword(s):  
Two Dimensional ◽  
Exonuclease Iii ◽  
Exo Iii ◽  
Recycling Amplification ◽  
Fluorescence Quencher

Herein a rapid and sensitive fluorometric bioanalysis platform for mercury(II) (Hg2+) detection was innovatively developed using ultrathin two-dimensional MXenes (Ti3C2) as fluorescence quencher and Hg2+-induced exonuclease III (Exo III)-assisted target...

Genomic footprinting detects factors bound to major late and IVa2 promoters in adenovirus-infected HeLa cells

1988 ◽  
Vol 8 (4) ◽  
pp. 1534-1539
Author(s):  
G Albrecht ◽  
B Devaux ◽  
C Kedinger
Keyword(s):  
Protein Interactions ◽  
Adenovirus Type ◽  
Nuclease Activity ◽  
Dnase I ◽  
Promoter Activation ◽  
Promoter Elements ◽  
Dnase I Footprinting ◽  
Infected Cells ◽  
Adenovirus Type 5

We used DNase I footprinting assays on nuclei isolated from adenovirus-infected cells to examine the nucleoprotein configuration of a 250-base-pair segment which encompasses the adenovirus type 5 major late (ML) and IVa2 promoters. At 12 and 20 h postinfection (p.i.), fine DNase I digestion mapping of wild-type adenovirus-infected cells revealed specific sequences protected from digestion which corresponded to promoter elements required for expression of the ML gene in vivo. At 12 h p.i., a G+C-rich region which lies upstream of the IVa2 cap site and is important for maximal IVa2 activity was also found masked to nuclease activity. At 20 h p.i., however, this element became more sensitive to nuclease attack, while the ML promoter elements stayed protected. No major changes in DNA-protein interactions were detected in the region spanning the ML and IVa2 cap sites upon promoter activation, suggesting that the binding properties of the cognate factors for this region are not modified during the process.

Label-free DNA detection based on oligonucleotide-stabilized silver nanoclusters and exonuclease III-catalyzed target recycling amplification

2014 ◽  
Vol 6 (15) ◽  
pp. 6082-6087 ◽  
Author(s):  
Hui Ma ◽  
Wei Wei ◽  
Qian Lu ◽  
Zhixin Zhou ◽  
Henan Li ◽  
...  
Keyword(s):  
High Sensitivity ◽  
Silver Nanoclusters ◽  
Label Free ◽  
Exonuclease Iii ◽  
Target Recycling ◽  
Free Dna ◽  
Sensitivity And Selectivity ◽  
Exo Iii ◽  
Ag Ncs ◽  
Recycling Amplification

A label-free DNA biosensor with high sensitivity and selectivity is constructed by using DNA–Ag NCs and Exo III-catalyzed target recycling amplification.

Target-activated cascaded digestion amplification of exonuclease III aided signal-on and ultrasensitive fluorescence detection of ATP

2018 ◽  
Vol 42 (5) ◽  
pp. 3534-3540 ◽  
Author(s):  
Xueqi Leng ◽  
Rongguo Li ◽  
Yu Wang ◽  
Yunping Wu ◽  
Yuqin Tu ◽  
...  
Keyword(s):  
Adenosine Triphosphate ◽  
Fluorescence Detection ◽  
Fluorescence Sensing ◽  
Exonuclease Iii ◽  
One Step ◽  
Exo Iii

In this work, a rapid, one-step and ultrasensitive signal-on fluorescence sensing for the detection of adenosine triphosphate (ATP) based on target-activated cascaded digestion amplification with Exo III aid was developed.

A crosslinked and ribosylated actin trimer does not interact productively with myosin

2019 ◽  
Vol 97 (2) ◽  
pp. 140-147 ◽  
Author(s):  
Navneet Sidhu ◽  
John F. Dawson
Keyword(s):  
Binding Site ◽  
Protein Interactions ◽  
Binding Proteins ◽  
Binding Protein ◽  
Dnase I ◽  
Protein Protein Interactions ◽  
Adp Ribosylation ◽  
Myosin Binding ◽  
Pointed End

A purified F-actin-derived actin trimer that interacts with end-binding proteins did not activate or bind the side-binding protein myosin under rigor conditions. Remodeling of the actin trimer by the binding of gelsolin did not rescue myosin binding, nor did the use of different means of inhibiting the polymerization of the trimer. Our results demonstrate that ADP-ribosylation on all actin subunits of an F-actin-derived trimer inhibits myosin binding and that the binding of DNase-I to the pointed end subunits of a crosslinked trimer also remodels the myosin binding site. Taken together, this work highlights the need for a careful balance between modification of actin subunits and maintaining protein–protein interactions to produce a physiologically relevant short F-actin complex.

Assessing the performance of the MM/PBSA and MM/GBSA methods. 10. Impacts of enhanced sampling and variable dielectric model on protein–protein Interactions

2019 ◽  
Vol 21 (35) ◽  
pp. 18958-18969 ◽  
Author(s):  
Ercheng Wang ◽  
Gaoqi Weng ◽  
Huiyong Sun ◽  
Hongyan Du ◽  
Feng Zhu ◽  
...  
Keyword(s):  
Protein Folding ◽  
Protein Interactions ◽  
Conformational Transitions ◽  
Protein Recognition ◽  
Enhanced Sampling ◽  
Protein Protein Interactions ◽  
Dielectric Model

Enhanced sampling has been extensively used to capture the conformational transitions in protein folding, but it attracts much less attention in the studies of protein–protein recognition.

scholarly journals Turn-On Fluorescence Aptasensor on Magnetic Nanobeads for Aflatoxin M1 Detection Based on an Exonuclease III-Assisted Signal Amplification Strategy

Nanomaterials ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 104
Author(s):  
Fuyuan Zhang ◽  
Linyang Liu ◽  
Shengnan Ni ◽  
Jiankang Deng ◽  
Guo-Jun Liu ◽  
...  
Keyword(s):  
Signal Amplification ◽  
Aflatoxin M1 ◽  
Exonuclease Iii ◽  
Sensor Performance ◽  
Milk Samples ◽  
Increased Sensitivity ◽  
Exo Iii ◽  
Amplification Strategy ◽  
Optimized Conditions ◽  
Magnetic Nanobeads

In order to satisfy the need for sensitive detection of Aflatoxin M1 (AFM1), we constructed a simple and signal-on fluorescence aptasensor based on an autocatalytic Exonuclease III (Exo III)-assisted signal amplification strategy. In this sensor, the DNA hybridization on magnetic nanobeads could be triggered by the target AFM1, resulting in the release of a single-stranded DNA to induce an Exo III-assisted signal amplification, in which numerous G-quadruplex structures would be produced and then associated with the fluorescent dye to generate significantly amplified fluorescence signals resulting in the increased sensitivity. Under the optimized conditions, this aptasensor was able to detect AFM1 with a practical detection limit of 9.73 ng kg−1 in milk samples. Furthermore, the prepared sensor was successfully used for detection of AFM1 in the commercially available milk samples with the recovery percentages ranging from 80.13% to 108.67%. Also, the sensor performance was evaluated by the commercial immunoassay kit with satisfactory results.

Multi-signal amplification electrochemical DNA biosensor based on exonuclease III and tetraferrocene

2020 ◽  
Vol 8 (18) ◽  
pp. 4143-4150
Author(s):  
Zhaojiang Yin ◽  
Hanfeng Cui ◽  
Qingxia Shu ◽  
Chen Jin ◽  
Yan Lin ◽  
...  
Keyword(s):  
Benzoic Acid ◽  
Electrochemical Sensor ◽  
Signal Amplification ◽  
Dna Biosensor ◽  
Exonuclease Iii ◽  
Electrochemical Dna Biosensor ◽  
Exo Iii

A new and efficient signal marker, 3,5-bis(3,5-bisferrocenethoxybenzyloxy)benzoic acid, was firstly synthesized and used for labeling. Construct a homogeneous electrochemical sensor with Exo III assisting the target cycle.

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