Recent Efforts and Milestones for Simulating Nucleic Acid FRET Experiments through Computational Methods

Author(s):  
Sorin Muraru ◽  
Sebastian Muraru ◽  
Florentin Romeo Nitu ◽  
Mariana Ionita
2020 ◽  
Vol 27 (5) ◽  
pp. 370-384
Author(s):  
Hua Wan ◽  
Jian-ming Li ◽  
Huang Ding ◽  
Shuo-xin Lin ◽  
Shu-qin Tu ◽  
...  

: Understanding the interaction mechanism of proteins and nucleic acids is one of the most fundamental problems for genome editing with engineered nucleases. Due to some limitations of experimental investigations, computational methods have played an important role in obtaining the knowledge of protein-nucleic acid interaction. Over the past few years, dozens of computational tools have been used for identification of nucleic acid binding site for site-specific proteins and design of site-specific nucleases because of their significant advantages in genome editing. Here, we review existing widely-used computational tools for target prediction of site-specific proteins as well as off-target prediction of site-specific nucleases. This article provides a list of on-line prediction tools according to their features followed by the description of computational methods used by these tools, which range from various sequence mapping algorithms (like Bowtie, FetchGWI and BLAST) to different machine learning methods (such as Support Vector Machine, hidden Markov models, Random Forest, elastic network and deep neural networks). We also make suggestions on the further development in improving the accuracy of prediction methods. This survey will provide a reference guide for computational biologists working in the field of genome editing.


Author(s):  
W. Bernard

In comparison to many other fields of ultrastructural research in Cell Biology, the successful exploration of genes and gene activity with the electron microscope in higher organisms is a late conquest. Nucleic acid molecules of Prokaryotes could be successfully visualized already since the early sixties, thanks to the Kleinschmidt spreading technique - and much basic information was obtained concerning the shape, length, molecular weight of viral, mitochondrial and chloroplast nucleic acid. Later, additonal methods revealed denaturation profiles, distinction between single and double strandedness and the use of heteroduplexes-led to gene mapping of relatively simple systems carried out in close connection with other methods of molecular genetics.


Author(s):  
Manfred E. Bayer

The first step in the infection of a bacterium by a virus consists of a collision between cell and bacteriophage. The presence of virus-specific receptors on the cell surface will trigger a number of events leading eventually to release of the phage nucleic acid. The execution of the various "steps" in the infection process varies from one virus-type to the other, depending on the anatomy of the virus. Small viruses like ØX 174 and MS2 adsorb directly with their capsid to the bacterial receptors, while other phages possess attachment organelles of varying complexity. In bacteriophages T3 (Fig. 1) and T7 the small conical processes of their heads point toward the adsorption site; a welldefined baseplate is attached to the head of P22; heads without baseplates are not infective.


Author(s):  
Dimitrij Lang

The success of the protein monolayer technique for electron microscopy of individual DNA molecules is based on the prevention of aggregation and orientation of the molecules during drying on specimen grids. DNA adsorbs first to a surface-denatured, insoluble cytochrome c monolayer which is then transferred to grids, without major distortion, by touching. Fig. 1 shows three basic procedures which, modified or not, permit the study of various important properties of nucleic acids, either in concert with other methods or exclusively:1) Molecular weights relative to DNA standards as well as number distributions of molecular weights can be obtained from contour length measurements with a sample standard deviation between 1 and 4%.


Author(s):  
Stephen D. Jett

The electrophoresis gel mobility shift assay is a popular method for the study of protein-nucleic acid interactions. The binding of proteins to DNA is characterized by a reduction in the electrophoretic mobility of the nucleic acid. Binding affinity, stoichiometry, and kinetics can be obtained from such assays; however, it is often desirable to image the various species in the gel bands using TEM. Present methods for isolation of nucleoproteins from gel bands are inefficient and often destroy the native structure of the complexes. We have developed a technique, called “snapshot blotting,” by which nucleic acids and nucleoprotein complexes in electrophoresis gels can be electrophoretically transferred directly onto carbon-coated grids for TEM imaging.


Author(s):  
B.A. Hamkalo ◽  
S. Narayanswami ◽  
A.P. Kausch

The availability of nonradioactive methods to label nucleic acids an the resultant rapid and greater sensitivity of detection has catapulted the technique of in situ hybridization to become the method of choice to locate of specific DNA and RNA sequences on chromosomes and in whole cells in cytological preparations in many areas of biology. It is being applied to problems of fundamental interest to basic cell and molecular biologists such as the organization of the interphase nucleus in the context of putative functional domains; it is making major contributions to genome mapping efforts; and it is being applied to the analysis of clinical specimens. Although fluorescence detection of nucleic acid hybrids is routinely used, certain questions require greater resolution. For example, very closely linked sequences may not be separable using fluorescence; the precise location of sequences with respect to chromosome structures may be below the resolution of light microscopy(LM); and the relative positions of sequences on very small chromosomes may not be feasible.


Author(s):  
M. Boublik ◽  
V. Mandiyan ◽  
S. Tumminia ◽  
J.F. Hainfeld ◽  
J.S. Wall

Success in protein-free deposition of native nucleic acid molecules from solutions of selected ionic conditions prompted attempts for high resolution imaging of nucleic acid interactions with proteins, not attainable by conventional EM. Since the nucleic acid molecules can be visualized in the dark-field STEM mode without contrasting by heavy atoms, the established linearity between scattering cross-section and molecular weight can be applied to the determination of their molecular mass (M) linear density (M/L), mass distribution and radius of gyration (RG). Determination of these parameters promotes electron microscopic imaging of biological macromolecules by STEM to a quantitative analytical level. This technique is applied to study the mechanism of 16S rRNA folding during the assembly process of the 30S ribosomal subunit of E. coli. The sequential addition of protein S4 which binds to the 5'end of the 16S rRNA and S8 and S15 which bind to the central domain of the molecule leads to a corresponding increase of mass and increased coiling of the 16S rRNA in the core particles. This increased compactness is evident from the decrease in RG values from 114Å to 91Å (in “ribosomal” buffer consisting of 10 mM Hepes pH 7.6, 60 mM KCl, 2 m Mg(OAc)2, 1 mM DTT). The binding of S20, S17 and S7 which interact with the 5'domain, the central domain and the 3'domain, respectively, continues the trend of mass increase. However, the RG values of the core particles exhibit a reverse trend, an increase to 108Å. In addition, the binding of S7 leads to the formation of a globular mass cluster with a diameter of about 115Å and a mass of ∽300 kDa. The rest of the mass, about 330 kDa, remains loosely coiled giving the particle a “medusa-like” appearance. These results provide direct evidence that 16S RNA undergoes significant structural reorganization during the 30S subunit assembly and show that its interactions with the six primary binding proteins are not sufficient for 16S rRNA coiling into particles resembling the native 30S subunit, contrary to what has been reported in the literature.


Author(s):  
D. P. Bazett-Jones ◽  
M. J. Hendzel

Structural analysis of combinations of nucleosomes and transcription factors on promoter and enhancer elements is necessary in order to understand the molecular mechanisms responsible for the regulation of transcription initiation. Such complexes are often not amenable to study by high resolution crystallographic techniques. We have been applying electron spectroscopic imaging (ESI) to specific problems in molecular biology related to transcription regulation. There are several advantages that this technique offers in studies of nucleoprotein complexes. First, an intermediate level of spatial resolution can be achieved because heavy atom contrast agents are not necessary. Second, mass and stoichiometric relationships of protein and nucleic acid can be estimated by phosphorus detection, an element in much higher proportions in nucleic acid than protein. Third, wrapping or bending of the DNA by the protein constituents can be observed by phosphorus mapping of the complexes. Even when ESI is used with high exposure of electrons to the specimen, important macromolecular information may be provided. For example, an image of the TATA binding protein (TBP) bound to DNA is shown in the Figure (top panel). It can be seen that the protein distorts the DNA away from itself and much of its mass sits off the DNA helix axis. Moreover, phosphorus and mass estimates demonstrate whether one or two TBP molecules interact with this particular promoter TATA sequence.


2021 ◽  
Vol 23 (1) ◽  
pp. 219-228
Author(s):  
Nabanita Saikia ◽  
Mohamed Taha ◽  
Ravindra Pandey

The rational design of self-assembled nanobio-molecular hybrids of peptide nucleic acids with single-wall nanotubes rely on understanding how biomolecules recognize and mediate intermolecular interactions with the nanomaterial's surface.


Sign in / Sign up

Export Citation Format

Share Document