Daniele Fabris

Albany State University, Georgia, United States

Are you Daniele Fabris?

Claim your profile

Publications (66)297.99 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Supported by high-throughput sequencing technologies, structure-specific nucleases are experiencing a renaissance as biochemical probes for genome-wide mapping of nucleic acid structure. This report explores the benefits and pitfalls of the application of Mung bean (Mb) and V1 nuclease, which attack specifically single- and double-stranded regions of nucleic acids, as possible structural probes to be employed in combination with MS detection. Both enzymes were found capable of operating in ammonium-based solutions that are preferred for high-resolution analysis by direct infusion electrospray ionization (ESI). Sequence analysis by tandem mass spectrometry (MS/MS) was performed to confirm mapping assignments and to resolve possible ambiguities arising from the concomitant formation of isobaric products with identical base composition and different sequences. The observed products grouped together into ladder-type series that facilitated their assignment to unique regions of the substrate, but revealed also a certain level of uncertainty in identifying the boundaries between paired and unpaired regions. Various experimental factors that are known to stabilize nucleic acid structure, such as higher ionic strength, presence of Mg(II), etc., increased the accuracy of cleavage information, but did not completely eliminate deviations from expected results. These observations suggest extreme caution in interpreting the results afforded by these types of reagents. Regardless of the analytical platform of choice, the results highlighted the need to repeat probing experiments under the most diverse possible conditions to recognize potential artifacts and to increase the level of confidence in the observed structural information.
    Journal of the American Society for Mass Spectrometry 05/2014; · 3.59 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The challenges posed by the analysis of mono-nucleotide mixtures by direct infusion electrospray ionization were examined in the context of recent advances of mass spectrometry (MS) technologies. In particular, we evaluated the merits of high-resolution mass analysis, multistep gas-phase dissociation, and ion mobility determinations for the characterization of species with very similar or identical elemental composition. The high resolving power afforded by a linear trap quadrupole-orbitrap allowed the complete differentiation of overlapping isotopic distributions produced by nucleotides that differed by a single mass unit. Resolving (12) C signals from nearly overlapped (13) C contributions provided the exact masses necessary to calculate matching elemental compositions for unambiguous formulae assignment. However, it was the ability to perform sequential steps of gas-phase dissociation (i.e. MS(n) -type analysis) that proved more valuable for discriminating between truly isobaric nucleotides, such as the AMP/dGMP and UMP/ΨMP couples, which were differentiated in the mixture from their unique fragmentation patterns. The identification of diagnostic fragments enabled the deconvolution of dissociation spectra containing the products of coexisting isobars that could not be individually isolated in the mass-selection step. Approaches based on ion mobility spectrometry-MS provided another dimension upon which isobaric nucleotides could be differentiated according to their distinctive mobility behaviors. Subtle structural variations, such as the different positions of an oxygen atom in AMP/dGMP or the glycosidic bond in UMP/ΨMP, produced detectable differences in the respective ion mobility profiles, which enabled the differentiation of the isobaric couples in the mixture. Parallel activation of all ions emerging from the ion mobility element provided an additional dimension for differentiating these analytes on the basis of both mobility and fragmentation properties. Copyright © 2013 John Wiley & Sons, Ltd.
    Biological Mass Spectrometry 06/2013; 48(6):703-12. · 3.41 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: In minimal RNA kissing complexes formed between hairpins with cognate GACG tetraloops, the two tertiary GC pairs are likely stabilized by the stacking of 5'-unpaired adenines at each end of the short helix. To test this hypothesis, we mutated the flanking adenines to various nucleosides and examined their effects on the kissing interaction. Electrospray ionization mass spectrometry was used to detect kissing dimers in a multi-equilibria mixture, whereas optical tweezers were applied to monitor the (un)folding trajectories of single RNA molecules. The experimental findings were rationalized by molecular dynamics simulations. Together, the results showed that the stacked adenines are indispensable for the tertiary interaction. By shielding the tertiary base pairs from solvent and reducing their fraying, the stacked adenines made terminal pairs act more like interior base pairs. The purine double-ring of adenine was essential for effective stacking, whereas additional functional groups modulated the stabilizing effects through varying hydrophobic and electrostatic forces. Furthermore, formation of the kissing complex was dominated by base pairing, whereas its dissociation was significantly influenced by the flanking bases. Together, these findings indicate that unpaired flanking nucleotides play essential roles in the formation of otherwise unstable two-base-pair RNA tertiary interactions.
    Journal of the American Chemical Society 03/2013; · 10.68 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: A stable RNA helix requires at least three base pairs. Surprisingly, a tertiary kissing complex formed between two GACG hairpin loops contains only two GC pairs. In the NMR structure of this complex, the two flanking adenosines stack on the kissing GC pair. This observation raised a possibility that the 5’-dangling adenines contribute to the formation and stability of the kissing interaction. To test this hypothesis, we took a two-pronged approach to examine the effects of various mutational and chemical modifications of the flanking adenosines on the folding of the kissing complex. Using mass spectrometry, we studied formation of kissing dimers formed by different hairpins. Using optical tweezers, we monitored mechanical unfolding of intramolecular kissing complex at single-molecule level. In both experiments, replacing adenine with uridine abolished the kissing interaction, suggesting that a minimal kissing complex must contain two GC pairs flanked by inter-strand stacking adenines. The stabilizing effect by the adenines can be explained by the fact that the stacking purine nucleobases shield the hydrogen bonds of the adjacent GC pairs, preventing them from fraying. Unlike in the context of secondary structure, the 5’-unpaired adenines in the tertiary structure are structurally constrained in a way that allows for effective stacking onto the adjacent base pairs.
    Biophysical Journal 01/2013; 104(2):411-. · 3.67 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Caught in the oxirane: Naphthalene diimides conjugated to a quinone methide and an oxirane have been synthesized and investigated as selective DNA G-quadruplex alkylating agents. The oxirane derivative generates a stable adduct with a G-quadruplex and shows selective alkylation of the loop adenines, as illustrated.
    Chemistry - A European Journal 12/2012; · 5.93 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Non-ergodic as well as ergodic activation methods are capable of maintaining the integrity of base pairs during the top-down analysis of nucleic acids. Here, we investigate the significance of this characteristic in the investigation of higher-order structures of increasing complexity. We show that cognate fragments produced by typical backbone cleavages may not be always detected as separate sequence ions, but rather as individual products that remain associated through mutual pairing contacts. This effect translates into unintended masking of cleavage events that take place in double-stranded regions, thus leading to the preferential detection of fragments originating from unpaired regions. Such effect is determined by the stability of the weak non-covalent association between complementary stretches, which is affected by base composition, length of the double-stranded structure, and charge of the precursor ion selected for analysis. Although such effect may prevent the achievement of full sequence coverage for primary structure determination, it may provide the key to correctly differentiate double- versus single-stranded regions, in what could be defined as gas-phase footprinting experiments. In light of the critical role played by base pairs in defining the higher-order structure of nucleic acids, these approaches will be expected to support an increased utilization of mass spectrometry for the investigation of nucleic acid structure and dynamics.
    International Journal of Mass Spectrometry 02/2012; 312:155-162. · 2.14 Impact Factor
  • Source
    D Fabris
    [Show abstract] [Hide abstract]
    ABSTRACT: Alternative approaches complementing the existing technologies for analysis of nucleic acids and their assemblies are necessary to take on the new challenges posed by the postgenomic era. The versatility of MS in biopolymer analysis and its ability to reach beyond sequence information are the basis of ever expanding applications aimed at the elucidation of nucleic acid structure-function relationships. This Feature summarizes the current state of MS-based approaches devised to overcome the limitations of traditional techniques and to advance different facets of nucleic acids research.
    Analytical Chemistry 06/2011; 83(15):5810-6. · 5.82 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Since its inception in 1994, The RNA Modification Database (RNAMDB, http://rna-mdb.cas.albany.edu/RNAmods/) has served as a focal point for information pertaining to naturally occurring RNA modifications. In its current state, the database employs an easy-to-use, searchable interface for obtaining detailed data on the 109 currently known RNA modifications. Each entry provides the chemical structure, common name and symbol, elemental composition and mass, CA registry numbers and index name, phylogenetic source, type of RNA species in which it is found, and references to the first reported structure determination and synthesis. Though newly transferred in its entirety to The RNA Institute, the RNAMDB continues to grow with two notable additions, agmatidine and 8-methyladenosine, appended in the last year. The RNA Modification Database is staying up-to-date with significant improvements being prepared for inclusion within the next year and the following year. The expanded future role of The RNA Modification Database will be to serve as a primary information portal for researchers across the entire spectrum of RNA-related research.
    Nucleic Acids Research 11/2010; 39(Database issue):D195-201. · 8.81 Impact Factor
  • Source
    Daniele Fabris, Eizadora T Yu
    [Show abstract] [Hide abstract]
    ABSTRACT: Chemical probing represents a very versatile alternative for studying the structure and dynamics of substrates that are intractable by established high-resolution techniques. The implementation of MS-based strategies for the characterization of probing products has not only extended the range of applicability to virtually all types of biopolymers but has also paved the way for the introduction of new reagents that would not have been viable with traditional analytical platforms. As the availability of probing data is steadily increasing on the wings of the development of dedicated interpretation aids, powerful computational approaches have been explored to enable the effective utilization of such information to generate valid molecular models. This combination of factors has contributed to making the possibility of obtaining actual 3D structures by MS-based technologies (MS3D) a reality. Although approaches for achieving structure determination of unknown targets or assessing the dynamics of known structures may share similar reagents and development trajectories, they clearly involve distinctive experimental strategies, analytical concerns and interpretation paradigms. This Perspective offers a commentary on methods aimed at obtaining distance constraints for the modeling of full-fledged structures while highlighting common elements, salient distinctions and complementary capabilities exhibited by methods used in dynamics studies. We discuss critical factors to be addressed for completing effective structural determinations and expose possible pitfalls of chemical methods. We survey programs developed for facilitating the interpretation of experimental data and discuss possible computational strategies for translating sparse spatial constraints into all-atom models. Examples are provided to illustrate how the concerted application of very diverse probing techniques can lead to the solution of actual biological systems.
    Biological Mass Spectrometry 08/2010; 45(8):841-60. · 3.41 Impact Factor
  • Source
    D Fabris, P A Limbach
    Journal of the American Society for Mass Spectrometry 03/2010; 21(7):R1-4. · 3.59 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Although polypurine tract (PPT)-primed initiation of plus-strand DNA synthesis in retroviruses and LTR-containing retrotransposons can be accurately duplicated, the molecular details underlying this concerted series of events remain largely unknown. Importantly, the PPT 3' terminus must be accommodated by ribonuclease H (RNase H) and DNA polymerase catalytic centers situated at either terminus of the cognate reverse transcriptase (RT), and in the case of the HIV-1 enzyme, ∼70Å apart. Communication between RT and the RNA/DNA hybrid therefore appears necessary to promote these events. The crystal structure of the HIV-1 RT/PPT complex, while informative, positions the RNase H active site several bases pairs from the PPT/U3 junction, and thus provides limited information on cleavage specificity. To fill the gap between biochemical and crystallographic approaches, we review a multidisciplinary approach combining chemical probing, mass spectrometry, NMR spectroscopy and single molecule spectroscopy. Our studies also indicate that nonnucleoside RT inhibitors affect enzyme orientation, suggesting initiation of plus-strand DNA synthesis as a potential therapeutic target.
    Viruses 12/2009; 1(3):657-77. · 2.51 Impact Factor
  • Source
    Daniele Fabris
    [Show abstract] [Hide abstract]
    ABSTRACT: The advances of mass spectrometry in the analysis of nucleic acids have tracked very closely the exciting developments of instrumentation and ancillary technologies, which have taken place over the years. However, their diffusion in the broader life sciences community has been and will be linked to the ever evolving focus of biomedical research and its changing demands. Before the completion of the Human Genome Project, great emphasis was placed on sequencing technologies that could help accomplish this project of exceptional scale. After the publication of the human genome, the emphasis switched toward techniques dedicated to the exploration of sequences not coding for actual protein products, which amount to the vast majority of transcribed elements. The broad range of capabilities offered by mass spectrometry is rapidly advancing this platform to the forefront of the technologies employed for the structure-function investigation of these noncoding elements. Increasing focus on the characterization of functional assemblies and their specific interactions has prompted a re-evaluation of what has been traditionally construed as nucleic acid analysis by mass spectrometry. Inspired by the accelerating expansion of the broader field of nucleic acid research, new applications to fundamental biological studies and drug discovery will help redefine the evolving role of MS-analysis of nucleic acids in the post-genomics age.
    Journal of the American Society for Mass Spectrometry 09/2009; 21(1):1-13. · 3.59 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: During (−)-strand DNA synthesis in retroviruses and Saccharomyces cerevisiae LTR retrotransposons, a purine rich region of the RNA template, known as the polypurine tract (PPT), is resistant to RNase H-mediated hydrolysis and subsequently serves as a primer for (+)-strand, DNA-dependent DNA synthesis. Although HIV-1 and Ty3 PPT sequences share no sequence similarity beyond the fact that both include runs of purine ribonucleotides, it has been suggested that these PPTs are processed by their cognate reverse transcriptases (RTs) through a common molecular mechanism. Here, we have used the aminoglycoside neomycin B (NB) to examine which structural features of the Ty3 PPT contribute to specific recognition and processing by its cognate RT. Using high-resolution NMR, direct infusion FTICR mass spectrometry, and isothermal titration calorimetry, we show that NB binds preferentially and selectively adjacent to the Ty3 3′ PPT−U3 cleavage junction and in an upstream 5′ region where the thumb subdomain of Ty3 RT putatively grips the substrate. Regions highlighted by NB on the Ty3 PPT are similar to those previously identified on the HIV-1 PPT sequence that are implicated as contact points for substrate binding by its RT. Our findings thus support the notion that common structural features of lentiviral and LTR-retrotransposon PPTs facilitate the interaction with their cognate RT.
    Biochemistry 07/2009; 48(29). · 3.38 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) has gained popularity as a facile method of examining RNA structure both in vitro and in vivo, exploiting accessibility of the ribose 2'-OH to acylation by N-methylisatoic anhydride (NMIA) in unpaired or flexible configurations. Subsequent primer extension terminates at the site of chemical modification, and these products are fractionated by high-resolution gel electrophoresis. When applying SHAPE to investigate structural features associated with the wild-type and analog-substituted polypurine tract (PPT)-containing RNA/DNA hybrids, their size (20-25 base pairs) rendered primer extension impractical. As an alternative method of detection, we reasoned that chemical modification could be combined with tandem mass spectrometry, relying on the mass increment of RNA fragments containing the NMIA adduct (M(r) = 133 Da). Using this approach, we demonstrate both specific modification of the HIV-1 PPT RNA primer and variations in its acylation pattern induced by replacing template nucleotides with a non-hydrogen-bonding thymine isostere. Our selective 2'-hydroxyl acylation analyzed by mass spectrometry strategy (SHAMS) should find utility when examining the structure of small RNA fragments or RNA/DNA hybrids where primer extension cannot be performed.
    RNA 07/2009; 15(8):1605-13. · 5.09 Impact Factor
  • Source
    Alexei Gapeev, Alberto Berton, Daniele Fabris
    [Show abstract] [Hide abstract]
    ABSTRACT: The hypothesis that direct determination of electrospray current would provide a viable method for maintaining spray stability to enable optimal nanospray analysis was tested by building a feedback apparatus capable of reading the current and readjusting the emitter voltage in real time. The apparatus consists of a current-sensing circuit that reads the voltage drop across a resistor located between the high-voltage power supply and the nanospray emitter. A low voltage proportional to the observed current is generated and sent to a data acquisition card. The information is used by a proportional-derivative-integral (PID) algorithm to calculate the magnitude of a low-voltage signal that is used to control the power supply output. Any variation of current across the sensing resistor is thus counteracted by an opposite-direction variation of the high voltage applied to the nanospray emitter. In this way, the apparatus adjusts the emitter voltage to achieve a preset value of current, which it strives to maintain over time in spite of any possible variation of the parameters influencing the spray regime. Preliminary results have shown that the feedback apparatus is capable of establishing and maintaining stable spray for samples that are usually considered challenging in traditional voltage-controlled analysis, such as those consisting of nucleic acid solutions with high salt loads. For these types of samples, the total ion count recorded in current-controlled mode was significantly more stable than that observed in voltage-controlled mode. At the same time, overall signal intensities and signal-to-noise ratios were also significantly improved. Setting the target nanospray current to a predefined value and letting the apparatus reach the target without operator intervention enabled the acquisition of viable data from solutions containing up to 2.5 M ammonium acetate, which are ordinarily difficult by traditional manual tuning. A deeper understanding of the current-voltage relationships for samples of very different compositions is expected to enable one not only to predict the target current that should be used for a certain analysis, but also to devise algorithms to change such target as a function of predictable variations of sample properties and analytical conditions. This will allow for optimal performance to be maintained during on-line gradient chromatography in which the nature of the sprayed solution may vary very widely during the course of the analysis.
    Journal of the American Society for Mass Spectrometry 04/2009; 20(7):1334-41. · 3.59 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The nature of specific RNA-RNA and protein-RNA interactions involved in the process of genome dimerization and isomerization in HIV-1, which is mediated in vitro by stemloop 1 (SL1) of the packaging signal and by the nucleocapsid (NC) domain of the viral Gag polyprotein, was investigated by using archetypical nucleic acid ligands as noncovalent probes. Small-molecule ligands make contact with their target substrates through complex combinations of H-bonds, salt bridges, and hydrophobic interactions. Therefore, their binding patterns assessed by electrospray ionization mass spectrometry can provide valuable insights into the factors determining specific recognition between species involved in biopolymer assemblies. In the case of SL1, dimerization and isomerization create unique structural features capable of sustaining stable interactions with classic nucleic acid ligands. The binding modes exhibited by intercalators and minor groove binders were adversely affected by the significant distortion of the duplex formed by palindrome annealing in the kissing-loop (KL) dimer, whereas the modes observed for the corresponding extended duplex (ED) confirmed a more regular helical structure. Consistent with the ability to establish electrostatic interactions with highly negative pockets typical of helix anomalies, polycationic aminoglycosides bound to the stem-bulge motif conserved in all SL1 conformers, to the unpaired nucleotides located at the hinge between kissing hairpins in KL, and to the exposed bases flanking the palindrome duplex in ED. The patterns afforded by intercalators and minor groove binders did not display detectable variations when the corresponding NC-SL1 complexes were submitted to probing. In contrast, aminoglycosides displayed the ability to compete with the protein for overlapping sites, producing opposite effects on the isomerization process. Indeed, displacing NC from the stem-bulges of the KL dimer induced inhibition of stem melting and decreased the efficiency of isomerization. Competition for the hinge region, instead, eliminated the NC stabilization of a grip motif formed by nucleobases of opposite strands, thus facilitating the strand-exchange required for isomerization. These noncovalent probes provided further evidence that the structural context of the actual binding sites has significant influence on the chaperone activities of NC, which should be taken in account when developing potential drug candidates aimed at disrupting genome dimerization and isomerization in HIV-1. © 2008 Wiley Periodicals, Inc. Biopolymers 91: 283–296, 2009.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com
    Biopolymers 03/2009; 91(4):283 - 296. · 2.88 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The combination of chemical probing and high-resolution mass spectrometry constitutes a powerful alternative for the structural elucidation of biomolecules possessing unfavorable size, solubility, and flexibility. We have developed nested Arg-specific bifunctional crosslinkers to obtain complementary information to typical Cys- and Lys-specific reagents available on the market. The structures of 1,4-phenyl-diglyoxal (PDG) and 4,4'-biphenyl-diglyoxal (BDG) include two identical 1,2-dicarbonyl functions capable of reacting with the guanido group of Arg residues in proteins, as well as the base-pairing face of guanine in nucleic acids. The reactive functions are separated by modular spacers consisting of one or two benzene rings, which confer greater rigidity to the crosslinker structure than it is afforded by typical aliphatic spacers. Analysis by electrospray ionization (ESI) Fourier transform ion cyclotron resonance (FTICR) mass spectrometry has shown that the probes provide both mono- and bifunctional products with model protein substrates, which are stabilized by the formation of diester derivatives in the presence of borate buffer. The identification of crosslinked sites was accomplished by employing complementary proteolytic procedures and peptide mapping by ESI-FTICR. The results showed excellent correlation with the solvent accessibility and structural context of susceptible residues, and highlighted the significance of possible dynamic effects in determining the outcome of crosslinking reactions. The application of nested reagents with different spacing has provided a new tool for experimentally recognizing flexible regions that may be involved in prominent dynamics in solution. The development of new bifunctional crosslinkers with diverse target specificity and different bridging spans is expected to facilitate the structure elucidation of progressively larger biomolecular assemblies by increasing the number and diversity of spatial constraints available for triangulating the position of crosslinked structures in the three dimensions.
    Analytica chimica acta 11/2008; 627(1):117-28. · 4.31 Impact Factor
  • Source
    Kevin B Turner, Sarah A Monti, Daniele Fabris
    [Show abstract] [Hide abstract]
    ABSTRACT: A rare example of ion/ion reaction between species of like polarity was shown to take place during the transfer of metal cations from nucleic acid substrates to chelating agents in the gas phase. Gaseous anionic reactants were generated from separate solutions of analyte and chelator by using a dual nanospray setup. The respective multiply charged ions shared the same path and were allowed to react for a predetermined interval in an rf-only hexapole before high-resolution analysis by Fourier transform ion cyclotron resonance (FTICR) mass spectrometry. Efficient transfer of sodium and magnesium ions was readily observed with significant reduction of the nonspecific adducts that are typically associated with decreased sensitivity and resolution in the analysis of nucleic acid samples. Metal cations were abstracted from the initial analyte without being replaced by protons, in a process that was clearly dependent on the concentration of chelator in the auxiliary emitter and on the time spent by the reactants in the hexapole element. A survey of the properties of selected anionic chelators showed that their known affinity for a target cation in solution was more critical than their maximum anionic charge in determining the outcome of the transfer process. The analysis of selected assemblies requiring divalent cations to preserve their structural integrity and functional properties demonstrated that ion/ion reactions were clearly capable of discriminating between nonspecific interactions and specific coordination based on transfer susceptibility. These examples demonstrated that the ability to selectively eliminate nonspecific adducts in the gas phase, after the desolvation process is complete, offers a unique opportunity for studying specific metal binding in biological systems without resorting to separation procedures that may adversely affect the position of binding equilibria in solution and disrupt the assemblies under investigation.
    Journal of the American Chemical Society 10/2008; 130(40):13353-63. · 10.68 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Modern biomedical research is evolving with the rapid growth of diverse data types, biophysical characterization methods, computational tools and extensive collaboration among researchers spanning various communities and having complementary backgrounds and expertise. Collaborating researchers are increasingly dependent on shared data and tools made available by other investigators with common interests, thus forming communities that transcend the traditional boundaries of the single research laboratory or institution. Barriers, however, remain to the formation of these virtual communities, usually due to the steep learning curve associated with becoming familiar with new tools, or with the difficulties associated with transferring data between tools. Recognizing the need for shared reference data and analysis tools, we are developing an integrated knowledge environment that supports productive interactions among researchers. Here we report on our current collaborative environment, which focuses on bringing together structural biologists working in the area of mass spectrometric based methods for the analysis of tertiary and quaternary macromolecular structures (MS3D) called the Collaboratory for MS3D (C-MS3D). C-MS3D is a Web-portal designed to provide collaborators with a shared work environment that integrates data storage and management with data analysis tools. Files are stored and archived along with pertinent meta data in such a way as to allow file handling to be tracked (data provenance) and data files to be searched using keywords and modification dates. While at this time the portal is designed around a specific application, the shared work environment is a general approach to building collaborative work groups. The goal of this is to not only provide a common data sharing and archiving system, but also to assist in the building of new collaborations and to spur the development of new tools and technologies.
    Journal of Proteome Research 10/2008; 7(11):4848-57. · 5.06 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The structure of HIV-1 Psi-RNA has been elucidated by a concerted approach combining structural probes with mass spectrometric detection (MS3D), which is not affected by the size and crystallization properties of target biomolecules. Distance constraints from bifunctional cross-linkers provided the information required for assembling an all-atom model from the high-resolution coordinates of separate domains by triangulating their reciprocal placement in 3D space. The resulting structure revealed a compact cloverleaf morphology stabilized by a long-range tertiary interaction between the GNRA tetraloop of stemloop 4 (SL4) and the upper stem of stemloop 1 (SL1). The preservation of discrete stemloop structures ruled out the possibility that major rearrangements might produce a putative supersite with enhanced affinity for the nucleocapsid (NC) domain of the viral Gag polyprotein, which would drive genome recognition and packaging. The steric situation of single-stranded regions exposed on the cloverleaf structure offered a valid explanation for the stoichiometry exhibited by full-length Psi-RNA in the presence of NC. The participation of SL4 in a putative GNRA loop-receptor interaction provided further indications of the plasticity of this region of genomic RNA, which can also anneal with upstream sequences to stabilize alternative conformations of the 5' untranslated region (5'-UTR). Considering the ability to sustain specific NC binding, the multifaceted activities supported by the SL4 sequence suggest a mechanism by which Gag could actively participate in regulating the vital functions mediated by 5'-UTR. Substantiated by the 3D structure of Psi-RNA, the central role played by SL4 in specific RNA-RNA and protein-RNA interactions advances this domain as a primary target for possible therapeutic intervention.
    Proceedings of the National Academy of Sciences 09/2008; 105(34):12248-53. · 9.81 Impact Factor

Publication Stats

841 Citations
297.99 Total Impact Points

Institutions

  • 2011–2013
    • Albany State University
      • Division of Chemistry
      Georgia, United States
  • 1993–2010
    • University of Maryland, Baltimore County
      • Department of Chemistry and Biochemistry
      Baltimore, Maryland, United States
  • 2008
    • National Cancer Institute (USA)
      Maryland, United States
  • 2003–2005
    • University of Padova
      Padua, Veneto, Italy
  • 1996–2005
    • University of Maryland, Baltimore
      Baltimore, Maryland, United States