Hydration of r(UGGGGU) quadruplexes

For about 50 years it has been known that guanine-rich sequence can, under appropriate conditions, adopt a distinctive, four-stranded, helical fold known as a G-quadruplex. Interest in quadruplex folds has grown in recent years as evidence of their biological relevance has accumulated from both sequence analysis and function-specific assays. The folds are unusually stable and their formation appears to require close management to maintain cell health; regulatory failure correlates with genomic instability and a number of cancer phenotypes. This thesis examines, by x-ray crystallography, the solvent structure of a previously reported tetramolecular RNA quadruplex, UGGGGU stabilized by Sr2+ ions. Crystal forms of the octameric assembly formed by this sequence exhibit unusually strong diffraction and anomalous signal enabling the construction of reliable models to a resolution of 0.85 Å. The solvent structure confirms hydration patterns reported for other nucleic acid helical conformations and provides support for the greater stability of RNA quadruplexes relative to DNA. Novel features detected in the octameric RNA assembly include a new crystal form and evidence of multiple conformations, among which one leading to the formation of a well-hydrated internal cavity. Though solvent is generally acknowledged to play a fundamental role in nucleic acid structure, its characterization from diffraction data remains challenging. To assist with this task, the thesis investigates two novel additions to the crystallographic methods arsenal. The first is segmentation of electron density maps into Morse-Smale basins characterized by uniform gradient flow. The second is the use of level set Fast Marching methods to compute the full distance field defined by the molecular surface. Both techniques show promise, though additional work will be required to yield effective tools. Gradient-flow segmentation provides an unambiguous way to gather all map density associated with a modeled atom and enables the calculation of novel volumetric properties including total basin density. Distance field calculation provides a unified framework for combining molecular surface calculation with surface-related queries including pocket and cavity detection and solvent travel depth

Structural Variations and Solvent Structure of r(UGGGGU) Quadruplexes Stabilized by Sr(2+) Ions.

Guanine-rich sequences can, under appropriate conditions, adopt a distinctive, four-stranded, helical fold known as a G-quadruplex. Interest in quadruplex folds has grown in recent years as evidence of their biological relevance has accumulated from both sequence analysis and function-specific assays. The folds are unusually stable and their formation appears to require close management to maintain cell health; regulatory failure correlates with genomic instability and a number of cancer phenotypes. Biologically relevant quadruplex folds are anticipated to form transiently in mRNA and in single-stranded, unwound DNA. To elucidate factors, including bound solvent, that contribute to the stability of RNA quadruplexes, we examine, by X-ray crystallography and small-angle X-ray scattering, the structure of a previously reported tetramolecular quadruplex, UGGGGU stabilized by Sr(2+) ions. Crystal forms of the octameric assembly formed by this sequence exhibit unusually strong diffraction and anomalous signal enabling the construction of reliable models to a resolution of 0.88Å. The solvent structure confirms hydration patterns reported for other nucleic acid helical conformations and provides support for the greater stability of RNA quadruplexes relative to DNA. Novel features detected in the octameric RNA assembly include a new crystal form, evidence of multiple conformations and structural variations in the 3' U tetrad, including one that leads to the formation of a hydrated internal cavity

Structural Variations and Solvent Structure of r(UGGGGU) Quadruplexes Stabilized by Sr(2+) Ions.

Guanine-rich sequences can, under appropriate conditions, adopt a distinctive, four-stranded, helical fold known as a G-quadruplex. Interest in quadruplex folds has grown in recent years as evidence of their biological relevance has accumulated from both sequence analysis and function-specific assays. The folds are unusually stable and their formation appears to require close management to maintain cell health; regulatory failure correlates with genomic instability and a number of cancer phenotypes. Biologically relevant quadruplex folds are anticipated to form transiently in mRNA and in single-stranded, unwound DNA. To elucidate factors, including bound solvent, that contribute to the stability of RNA quadruplexes, we examine, by X-ray crystallography and small-angle X-ray scattering, the structure of a previously reported tetramolecular quadruplex, UGGGGU stabilized by Sr(2+) ions. Crystal forms of the octameric assembly formed by this sequence exhibit unusually strong diffraction and anomalous signal enabling the construction of reliable models to a resolution of 0.88Å. The solvent structure confirms hydration patterns reported for other nucleic acid helical conformations and provides support for the greater stability of RNA quadruplexes relative to DNA. Novel features detected in the octameric RNA assembly include a new crystal form, evidence of multiple conformations and structural variations in the 3' U tetrad, including one that leads to the formation of a hydrated internal cavity

Carotenoid stoichiometry in the LH2 crystal: No spectral evidence for the presence of the second molecule in the alpha/beta-apoprotein dimer

In this work we have investigated the carotenoid–protein interactions in LH2 complexes of <i>Rhodopseudomonas acidophila</i> both in “free in solution” mixed-micelles and in three-dimensional crystals by Raman spectroscopy in resonance with the carotenoid (Car) molecules. We show that the Car molecules when bound to their binding pockets show no significant differences when the complexes are “free in solution” or packed in crystalline arrays. Furthermore, there is no significant wavelength dependence in the Raman spectrum of the Car molecules of LH2. This indicates that there is only one Car configuration in LH2 and thus only one molecule per α/β-heterodimer

The UCSC SARS-CoV-2 Genome Browser.

BACKGROUND:: Researchers are generating molecular data pertaining to the SARS-CoV-2 RNA genome and its proteins at an unprecedented rate during the COVID-19 pandemic. As a result, there is a critical need for rapid and continuously updated access to the latest molecular data in a format in which all data can be quickly cross-referenced and compared. We adapted our genome browser visualization tool to the viral genome for this purpose. Molecular data, curated from published studies or from database submissions, are mapped to the viral genome and grouped together into “annotation tracks” where they can be visualized along the linear map of the viral genome sequence and programmatically downloaded in standard format for analysis. RESULTS:: The UCSC Genome Browser for SARS-CoV-2 (https://genome.ucsc.edu/covid19.html) provides continuously updated access to the mutations in the many thousands of SARS-CoV-2 genomes deposited in GISAID and the international nucleotide sequencing databases, displayed alongside phylogenetic trees. These data are augmented with alignments of bat, pangolin, and other animal and human coronavirus genomes, including per-base evolutionary rate analysis. All available annotations are cross-referenced on the virus genome, including those from major databases (PDB, RFAM, IEDB, UniProt) as well as up-to-date individual results from preprints. Annotated data include predicted and validated immune epitopes, promising antibodies, RT-PCR and sequencing primers, CRISPR guides (from research, diagnostics, vaccines, and therapies), and points of interaction between human and viral genes. As a community resource, any user can add manual annotations which are quality checked and shared publicly on the browser the next day. CONCLUSIONS:: We invite all investigators to contribute additional data and annotations to this resource to accelerate research and development activities globally. Contact us at [email protected] with data suggestions or requests for support for adding data. Rapid sharing of data will accelerate SARS-CoV-2 research, especially when researchers take time to integrate their data with those from other labs on a widely-used community browser platform with standardized machine-readable data formats, such as the SARS-CoV-2 Genome Browser

The UCSC Genome Browser database: 2021 update.

For more than two decades, the UCSC Genome Browser database (https://genome.ucsc.edu) has provided high-quality genomics data visualization and genome annotations to the research community. As the field of genomics grows and more data become available, new modes of display are required to accommodate new technologies. New features released this past year include a Hi-C heatmap display, a phased family trio display for VCF files, and various track visualization improvements. Striving to keep data up-to-date, new updates to gene annotations include GENCODE Genes, NCBI RefSeq Genes, and Ensembl Genes. New data tracks added for human and mouse genomes include the ENCODE registry of candidate cis-regulatory elements, promoters from the Eukaryotic Promoter Database, and NCBI RefSeq Select and Matched Annotation from NCBI and EMBL-EBI (MANE). Within weeks of learning about the outbreak of coronavirus, UCSC released a genome browser, with detailed annotation tracks, for the SARS-CoV-2 RNA reference assembly

The UCSC Genome Browser database: 2021 update.

For more than two decades, the UCSC Genome Browser database (https://genome.ucsc.edu) has provided high-quality genomics data visualization and genome annotations to the research community. As the field of genomics grows and more data become available, new modes of display are required to accommodate new technologies. New features released this past year include a Hi-C heatmap display, a phased family trio display for VCF files, and various track visualization improvements. Striving to keep data up-to-date, new updates to gene annotations include GENCODE Genes, NCBI RefSeq Genes, and Ensembl Genes. New data tracks added for human and mouse genomes include the ENCODE registry of candidate cis-regulatory elements, promoters from the Eukaryotic Promoter Database, and NCBI RefSeq Select and Matched Annotation from NCBI and EMBL-EBI (MANE). Within weeks of learning about the outbreak of coronavirus, UCSC released a genome browser, with detailed annotation tracks, for the SARS-CoV-2 RNA reference assembly