Single-Molecule Microscopy Meets Molecular Dynamics Simulations for Characterizing the Molecular Action of Proteins on DNA and in Liquid Condensates

Single Molecule ◽

Binding Proteins ◽

Dynamic Properties ◽

Dna Recombination ◽

Dna Binding Proteins ◽

Molecular Action ◽

DNA-binding proteins trigger various cellular functions and determine cellular fate. Before performing functions such as transcription, DNA repair, and DNA recombination, DNA-binding proteins need to search for and bind to their target sites in genomic DNA. Under evolutionary pressure, DNA-binding proteins have gained accurate and rapid target search and binding strategies that combine three-dimensional search in solution, one-dimensional sliding along DNA, hopping and jumping on DNA, and intersegmental transfer between two DNA molecules. These mechanisms can be achieved by the unique structural and dynamic properties of these proteins. Single-molecule fluorescence microscopy and molecular dynamics simulations have characterized the molecular actions of DNA-binding proteins in detail. Furthermore, these methodologies have begun to characterize liquid condensates induced by liquid-liquid phase separation, e.g., molecular principles of uptake and dynamics in droplets. This review discusses the molecular action of DNA-binding proteins on DNA and in liquid condensate based on the latest studies that mainly focused on the model protein p53.

Sequential eviction of crowded nucleoprotein complexes by the RecBCD molecular motor

10.1101/142224 ◽

2017 ◽

Author(s):

Tsuyoshi Terakawa ◽

Sy Redding ◽

Timothy D. Silverstein ◽

Eric C. Greene

Keyword(s):

Molecular Dynamics ◽

Nucleic Acid ◽

Dna Binding ◽

Single Molecule ◽

Motor Proteins ◽

Dna Binding Proteins ◽

Nucleoprotein Complexes ◽

Dynamics Simulations ◽

Crowded Environments

AbstractIn physiological settings, all nucleic acids motor proteins must travel along substrates that are crowded with other proteins. However, the physical basis for how motor proteins behave in these highly crowded environments remains unknown. Here we use real–time single molecule imaging, kinetic Monte Carlo simulations, and Molecular dynamics simulations to determine how the ATP-dependent translocase RecBCD travels along DNA occupied by tandem arrays of high affinity DNA–binding proteins. We demonstrate that RecBCD forces each protein into its nearest adjacent neighbor, causing rapid disruption of the underlying protein–nucleic acid interface. This mechanism is not simply the same way that RecBCD disrupts isolated nucleoprotein complexes on otherwise naked DNA. Instead, molecular crowding itself completely alters the mechanism by which RecBCD removes tightly bound protein obstacles from DNA.Significance statementChromosomes are crowded places, and any nucleic acid motor proteins that act upon DNA must function within these crowded environments. How crowded environments affect motor protein behaviors remains largely unexplored. Here, we use single molecule fluorescence microscopy visualize the ATP-dependent motor protein RecBCD as it travels along crowded DNA molecules bearing long tandem arrays of DNA-binding proteins. Our findings show that RecBCD can push through highly crowded protein arrays while evicting the proteins from DNA. Molecular dynamics simulations suggest that RecBCD forces the proteins into once another, causing rapid disruption of the protein-DNA interface. These findings may provide insights into how other types of motor proteins travel along crowded nucleic acids.

Methylation-targeted specificity of the DNA binding proteins R.DpnI and MeCP2 studied by molecular dynamics simulations

Journal of Molecular Modeling ◽

10.1007/s00894-017-3318-8 ◽

2017 ◽

Vol 23 (5) ◽

Cited By ~ 2

Author(s):

Siba Shanak ◽

Ozlem Ulucan ◽

Volkhard Helms

Keyword(s):

Molecular Dynamics ◽

Dna Binding ◽

Binding Proteins ◽

Dna Binding Proteins ◽

The HMGB chromatin protein Nhp6A can bypass obstacles when traveling on DNA

Nucleic Acids Research ◽

10.1093/nar/gkaa799 ◽

2020 ◽

Vol 48 (19) ◽

pp. 10820-10831

Author(s):

Kiyoto Kamagata ◽

Kana Ouchi ◽

Cheng Tan ◽

Eriko Mano ◽

Sridhar Mandali ◽

...

Keyword(s):

Dna Binding ◽

Single Molecule ◽

Binding Proteins ◽

Dna Binding Proteins ◽

Binding Modes ◽

Dna Structures ◽

Bacterial Proteins ◽

Associated Proteins ◽

Diffusion Mechanisms ◽

Abstract DNA binding proteins rapidly locate their specific DNA targets through a combination of 3D and 1D diffusion mechanisms, with the 1D search involving bidirectional sliding along DNA. However, even in nucleosome-free regions, chromosomes are highly decorated with associated proteins that may block sliding. Here we investigate the ability of the abundant chromatin-associated HMGB protein Nhp6A from Saccharomyces cerevisiae to travel along DNA in the presence of other architectural DNA binding proteins using single-molecule fluorescence microscopy. We observed that 1D diffusion by Nhp6A molecules is retarded by increasing densities of the bacterial proteins Fis and HU and by Nhp6A, indicating these structurally diverse proteins impede Nhp6A mobility on DNA. However, the average travel distances were larger than the average distances between neighboring proteins, implying Nhp6A is able to bypass each of these obstacles. Together with molecular dynamics simulations, our analyses suggest two binding modes: mobile molecules that can bypass barriers as they seek out DNA targets, and near stationary molecules that are associated with neighboring proteins or preferred DNA structures. The ability of mobile Nhp6A molecules to bypass different obstacles on DNA suggests they do not block 1D searches by other DNA binding proteins.

Regulation of DNA Metabolism by DNA-Binding Proteins Probed by Single Molecule Spectroscopy

10.21236/ada459264 ◽

2006 ◽

Author(s):

Andreas Hanke

Keyword(s):

Dna Binding ◽

Single Molecule ◽

Binding Proteins ◽

Dna Binding Proteins ◽

Single Molecule Spectroscopy ◽

Dna Metabolism

Dynamic properties of SARS-CoV and SARS-CoV-2 RNA-dependent RNA polymerases studied by molecular dynamics simulations

Chemical Physics Letters ◽

10.1016/j.cplett.2021.138819 ◽

2021 ◽

pp. 138819

Author(s):

Satoru G. Itoh ◽

Shoichi Tanimoto ◽

Hisashi Okumura

Keyword(s):

Molecular Dynamics ◽

Dynamic Properties ◽

Rna Polymerases ◽

Unraveling cGAS catalytic mechanism upon DNA activation through molecular dynamics simulations

Physical Chemistry Chemical Physics ◽

10.1039/d1cp00378j ◽

2021 ◽

Author(s):

Jordi Soler ◽

Pedro Paiva ◽

Maria Joao Joao Ramos ◽

Pedro Alexandrino Fernandes ◽

Marie Brut

Keyword(s):

Molecular Dynamics ◽

Dna Binding ◽

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...