scholarly journals DNA mismatches reveal widespread conformational penalties in protein-DNA recognition

2019 ◽  
Author(s):  
Ariel Afek ◽  
Honglue Shi ◽  
Atul Rangadurai ◽  
Harshit Sahay ◽  
Hashim M. Al-Hashimi ◽  
...  
Keyword(s):  
Dna Binding ◽  
Dna Sequence ◽  
Dna Recognition ◽  
Energetic Cost ◽  
Base Pairs ◽  
Structural Distortions ◽  
Dna Mismatches ◽  
Specific Affinity ◽  
Dynamics Simulations ◽  
High Throughput Assay

ABSTRACTTranscription-factor (TF) proteins recognize specific genomic sequences, despite an overwhelming excess of non-specific DNA, to regulate complex gene expression programs1–3. While there have been significant advances in understanding how DNA sequence and shape contribute to recognition, some fundamental aspects of protein-DNA binding remain poorly understood2,3. Many DNA-binding proteins induce changes in the DNA structure outside the intrinsic B-DNA envelope. How the energetic cost associated with distorting DNA contributes to recognition has proven difficult to study and measure experimentally because the distorted DNA structures exist as low-abundance conformations in the naked B-DNA ensemble4–10. Here, we use a novel high-throughput assay called SaMBA (Saturation Mismatch-Binding Assay) to investigate the role of DNA conformational penalties in TF-DNA recognition. The approach introduces mismatched base-pairs (i.e. mispairs) within TF binding sites to pre-induce a variety of DNA structural distortions much larger than those induced by changes in Watson-Crick sequence. Strikingly, while most mismatches either weakened TF binding (~70%) or had negligible effects (~20%), approximately 10% of mismatches increased binding and at least one mismatch was found that increased the binding affinity for each of 21 examined TFs. Mismatches also converted sites from the non-specific affinity range into specific sites, and high-affinity sites into “super-sites” stronger than any known canonical binding site. These findings reveal a complex binding landscape that cannot be explained based on DNA sequence alone. Analysis of crystal structures together with NMR and molecular dynamics simulations revealed that many of the mismatches that increase binding induce distortions similar to those induced by TF binding, thus pre-paying some of the energetic cost to deform the DNA. Our work indicates that conformational penalties are a major determinant of protein-DNA recognition, and reveals mechanisms by which mismatches can recruit TFs and thus modulate replication and repair activities in the cell11,12.

scholarly journals Rational design of a DNA sequence-specific modular protein tag by tuning the alkylation kinetics

2019 ◽  
Vol 10 (40) ◽  
pp. 9315-9325 ◽  
Author(s):  
Thang Minh Nguyen ◽  
Eiji Nakata ◽  
Zhengxiao Zhang ◽  
Masayuki Saimura ◽  
Huyen Dinh ◽  
...  
Keyword(s):  
Dna Binding ◽  
Kinetic Parameters ◽  
Dna Sequence ◽  
Rational Design ◽  
Design Principle ◽  
Dna Recognition ◽  
Protein Tag

A design principle for sequence-specific DNA modifiers driven by the specific DNA recognition was proposed based on the kinetic parameters for DNA binding and modification reactions.

scholarly journals DNA sequence context controls the binding and processivity of the T7 DNA primase

2018 ◽  
Author(s):  
Ariel Afek ◽  
Stefan Ilic ◽  
John Horton ◽  
David B. Lukatsky ◽  
Raluca Gordan ◽  
...  
Keyword(s):  
Dna Binding ◽  
Dna Sequence ◽  
Functional Activity ◽  
Dna Recognition ◽  
Unique Combination ◽  
Binding Assays ◽  
Sequence Context ◽  
Sequence Composition ◽  
Complex Landscape ◽  
Biochemical Analyses

SUMMARYPrimases are key enzymes involved in DNA replication. They act on single-stranded DNA, and catalyze the synthesis of short RNA primers used by DNA polymerases. Here, we investigate the DNA-binding and activity of the bacteriophage T7 primase using a new workflow called High-Throughput Primase Profiling (HTPP). Using a unique combination of high-throughput binding assays and biochemical analyses, HTPP reveals a complex landscape of binding specificity and functional activity for the T7 primase, determined by sequences flanking the primase recognition site. We identified specific features, such as G/T-rich flanks, which increase primase-DNA binding up to 10-fold and, surprisingly, also increase the length of newly formed RNA (up to 3-fold). To our knowledge, variability in primer length has not been reported for this primase. We expect that applying HTPP to additional enzymes will reveal new insights into the effects of DNA sequence composition on the DNA recognition and functional activity of primases.

scholarly journals The Small Subunit of M · AquI Is Responsible for Sequence-Specific DNA Recognition and Binding in the Absence of the Catalytic Domain

2003 ◽  
Vol 185 (4) ◽  
pp. 1284-1288 ◽  
Author(s):  
Hatice Pinarbasi ◽  
Ergun Pinarbasi ◽  
David P. Hornby
Keyword(s):  
Dna Binding ◽  
Dna Sequence ◽  
Dna Methyltransferase ◽  
Dna Recognition ◽  
Dna Methyltransferases ◽  
Binary Complex ◽  
Β Subunit ◽  
Α Subunit ◽  
Conserved Sequence ◽  
Covalent Complex

ABSTRACT AquI DNA methyltransferase (M · AquI) catalyzes the transfer of a methyl group from S-adenosyl-l-methionine to the C5 position of the outermost deoxycytidine base in the DNA sequence 5′-CCCGGG-3′. M · AquI is a heterodimer in which the polypeptide chain is separated at the junction between the two equivalent structural domains in the related enzyme M · HhaI. Recently, we reported the subcloning, overexpression, and purification of the subunits (α and β) of M · AquI separately. Here we describe the DNA binding properties of M · AquI. The results presented here indicate that the β subunit alone contains all of the information for sequence-specific DNA recognition and binding. The first step in the sequence-specific recognition of DNA by M · AquI involves the formation of binary complex with the target recognition domain in conjunction with conserved sequence motifs IX and X, found in all known C5 DNA methyltransferases, contained in the β subunit. The α subunit enhances the binding of the β subunit to DNA specifically and nonspecifically. It is likely that the addition of the α subunit to the β subunit stabilizes the conformation of the β subunit and thereby enhances its affinity for DNA indirectly. Addition of S-adenosyl-l-methionine and its analogues S-adenosyl-l-homocysteine and sinefungin enhances binding, but only in the presence of the α subunit. These compounds did not have any effect on DNA binding by the β subunit alone. Using a 30-mer oligodeoxynucleotide substrate containing 5-fluorodeoxycytidine (5-FdC), it was found that the β subunit alone did not form a covalent complex with its specific sequence in the absence or presence of S-adenosyl-l-methionine. However, the addition of the α subunit to the β subunit led to the formation of a covalent complex with specific DNA sequence containing 5-FdC.

scholarly journals A DNA Sequence Based Polymer Model for Chromatin Folding

2021 ◽  
Vol 22 (3) ◽  
pp. 1328
Author(s):  
Rui Zhou ◽  
Yi Qin Gao
Keyword(s):  
Dna Sequence ◽  
Cpg Island ◽  
Coarse Grained ◽  
Chromosome Territories ◽  
Base Pairs ◽  
Single Chromosome ◽  
Power Law Decay ◽  
Multiple Length Scales ◽  
Spatial Domains ◽  
Chromatin Folding

The recent development of sequencing technology and imaging methods has provided an unprecedented understanding of the inter-phase chromatin folding in mammalian nuclei. It was found that chromatin folds into topological-associated domains (TADs) of hundreds of kilo base pairs (kbps), and is further divided into spatially segregated compartments (A and B). The compartment B tends to be located near to the periphery or the nuclear center and interacts with other domains of compartments B, while compartment A tends to be located between compartment B and interacts inside the domains. These spatial domains are found to highly correlate with the mosaic CpG island (CGI) density. High CGI density corresponds to compartments A and small TADs, and vice versa. The variation of contact probability as a function of sequential distance roughly follows a power-law decay. Different chromosomes tend to segregate to occupy different chromosome territories. A model that can integrate these properties at multiple length scales and match many aspects is highly desired. Here, we report a DNA-sequence based coarse-grained block copolymer model that considers different interactions between blocks of different CGI density, interactions of TAD formation, as well as interactions between chromatin and the nuclear envelope. This model captures the various single-chromosome properties and partially reproduces the formation of chromosome territories.

Unraveling cGAS catalytic mechanism upon DNA activation through molecular dynamics simulations

2021 ◽  
Author(s):  
Jordi Soler ◽  
Pedro Paiva ◽  
Maria Joao Joao Ramos ◽  
Pedro Alexandrino Fernandes ◽  
Marie Brut
Keyword(s):  
Molecular Dynamics ◽  
Dna Binding ◽  
Molecular Dynamics Simulations ◽  
Cyclic Gmp ◽  
Catalytic Mechanism ◽  
System P ◽  
Dynamics Simulations ◽  
Cyclic Dinucleotide

Cyclic GMP-AMP Synthase (cGAS) is activated upon DNA binding and catalyzes the synthesis of 2’,3’-cGAMP from GTP and ATP. This cyclic dinucleotide is a messenger that triggers the autoimmune system...

scholarly journals Synthesis, Characterization, DNA Binding, and Photocleavage Activity of Oxorhenium (V) Complexes withα-Diimine and Quinoxaline Ligands

2010 ◽  
Vol 2010 ◽  
pp. 1-9 ◽  
Author(s):  
Christiana A. Mitsopoulou ◽  
Constantinos Dagas
Keyword(s):  
Cyclic Voltammetry ◽  
Dna Binding ◽  
Dna Cleavage ◽  
2D Nmr ◽  
Base Pairs ◽  
Binding Properties ◽  
Supercoiled Form ◽  
Dna Base ◽  
Dna Base Pairs ◽  
Dna Binding Properties

The complex [ReOCl3pq] (1) (where pq = 2-(2′pyridyl)quinoxaline) has been synthesized and fully characterized by UV-Vis, FTIR, 1 and 2D NMR, and cyclic voltammetry (CV). The DNA-binding properties of the complex1as well as of the compounds [ReOCl3bpy] (2), [ReOCl3phen] (3), and pq (4) were investigated by UV-spectrophotometric (melting curves), CV (cyclic voltammetry), and viscosity measurements. Experimental data suggest that complex1intercalates into the DNA base pairs. Upon irradiation, complex1was found to promote the cleavage of plasmid pBR 322 DNA from supercoiled form I to nicked form II. The mechanism of the DNA cleavage by complex1was also investigated.

scholarly journals On the dynamics of some small structural motifs in rRNA upon ligand binding

2008 ◽  
Vol 73 (1) ◽  
pp. 41-53
Author(s):  
Aleksandra Rakic ◽  
Petar Mitrasinovic
Keyword(s):  
Molecular Dynamics ◽  
Conformational Changes ◽  
Binding Sites ◽  
De Novo ◽  
Reference Structure ◽  
Base Pairs ◽  
Rna Sequences ◽  
Conformational Model ◽  
Thermodynamic Variables ◽  
Dynamics Simulations

The present study characterizes using molecular dynamics simulations the behavior of the GAA (1186-1188) hairpin triloops with their closing c-g base pairs in large ribonucleoligand complexes (PDB IDs: 1njn, 1nwy, 1jzx). The relative energies of the motifs in the complexes with respect to that in the reference structure (unbound form of rRNA; PDB ID: 1njp) display the trends that agree with those of the conformational parameters reported in a previous study1 utilizing the de novo pseudotorsional (?,?) approach. The RNA regions around the actual RNA-ligand contacts, which experience the most substantial conformational changes upon formation of the complexes were identified. The thermodynamic parameters, based on a two-state conformational model of RNA sequences containing 15, 21 and 27 nucleotides in the immediate vicinity of the particular binding sites, were evaluated. From a more structural standpoint, the strain of a triloop, being far from the specific contacts and interacting primarily with other parts of the ribosome, was established as a structural feature which conforms to the trend of the average values of the thermodynamic variables corresponding to the three motifs defined by the 15-, 21- and 27-nucleotide sequences. From a more functional standpoint, RNA-ligand recognition is suggested to be presumably dictated by the types of ligands in the complexes.

scholarly journals A plant DNA-binding protein that recognizes 5-methylcytosine residues.

1989 ◽  
Vol 9 (3) ◽  
pp. 1351-1356 ◽  
Author(s):  
D L Zhang ◽  
K C Ehrlich ◽  
P C Supakar ◽  
M Ehrlich
Keyword(s):  
Dna Binding ◽  
Dna Sequence ◽  
Base Pair ◽  
Binding Protein ◽  
Dna Binding Protein ◽  
Sequence Specificity ◽  
Dna Sequence Specificity ◽  
Plant Dna ◽  
Related Family ◽  
Nuclear Extracts

A novel, 5-methylcytosine-specific, DNA-binding protein, DBP-m, has been identified in nuclear extracts of peas. DBP-m specifically recognizes 5-methylcytosine residues in DNA without appreciable DNA sequence specificity, unlike a mammalian DNA-binding protein (MDBP), which recognizes 5-methylcytosine residues but only in a related family of 14-base-pair sequences.

Sign in / Sign up

Export Citation Format

Share Document