A5030180511- RNA and protein synthesis mechanisms
- RNA modifications and cancer
- DNA and Nucleic Acid Chemistry
- Marine Sponges and Natural Products
- Biochemical and Molecular Research
- Phytochemical Studies and Bioactivities
- Natural product bioactivities and synthesis
- Phytochemistry and Bioactive Compounds
- Sesquiterpenes and Asteraceae Studies
- Electron Spin Resonance Studies
- Galectins and Cancer Biology
- Bacterial Genetics and Biotechnology
- Glycosylation and Glycoproteins Research
- Traditional and Medicinal Uses of Annonaceae
- Metalloenzymes and iron-sulfur proteins
- RNA Research and Splicing
- Toxin Mechanisms and Immunotoxins
- Protein Structure and Dynamics
- Electrocatalysts for Energy Conversion
- Phytochemistry and Biological Activities
- Botanical Research and Chemistry
- Bacteriophages and microbial interactions
- Advanced biosensing and bioanalysis techniques
- Mass Spectrometry Techniques and Applications
- Advanced NMR Techniques and Applications
Cassia (United States)
2008-2024
Scripps Research Institute
2000-2010
Medical University of South Carolina
2004-2007
Bruker (United States)
2006
Masaryk University
2006
University of Toronto
2006
Palo Alto Research Center
2004
Genomics Institute of the Novartis Research Foundation
2004
Boston University
1995-2002
Williams College
1998
Radical S-adenosyl-l-methionine (SAM) enzymes comprise a vast superfamily catalyzing diverse reactions essential to all life through homolytic SAM cleavage liberate the highly reactive 5'-deoxyadenosyl radical (5'-dAdo·). Our recent observation of catalytically competent organometallic intermediate Ω that forms during reaction (RS) enzyme pyruvate formate-lyase activating-enzyme (PFL-AE) was therefore quite surprising, and led question its broad relevance in superfamily. We now show PFL-AE...
We report the fabrication of enthalpy arrays and their use to detect molecular interactions, including protein-ligand binding, enzymatic turnover, mitochondrial respiration. Enthalpy provide a universal assay methodology with no need for specific development such as fluorescent labeling or immobilization reagents, which can adversely affect interaction. Microscale technology enables 96-detector on large substrates. The reduction in scale results decreases both sample quantity measurement...
A general method for isotopic labeling of the purine base moiety nucleotides and RNA has been developed through biochemical pathway engineering in vitro. synthetic scheme was designed implemented utilizing recombinant enzymes from pentose phosphate de novo synthesis pathways, with regeneration folate, aspartate, glutamine, ATP, NADPH cofactors, a single-pot reaction. Syntheses proceeded quickly efficiently comparison to chemical methods isolated yields up 66% 13C-, 15N-enriched ATP GTP. The...
Enzymatic synthesis methods for the fluorinated 5'-triphosphate analogues 5F-UTP and 5F-CTP have been developed to facilitate 19F-labeling of RNAs biophysical studies. HIV-2 TAR were synthesized using these by in vitro transcription reactions T7 RNA polymerase. The uniform incorporation 5F-U or 5F-C into transcripts does not significantly alter structure thermodynamic stability. Fluorine observed homonuclear 19F−19F heteronuclear 19F−1H NOE experiments providing selective distance...
The use of stable isotope labeling has revolutionized NMR studies nucleic acids, and there is a need for methods incorporation specific labels to facilitate experiments applications. Enzymatic synthesis offers an efficient flexible means synthesize nucleoside triphosphates from variety commercially available specifically labeled precursors, permitting RNAs prepared by in vitro transcription. Here, we recapitulate de novo pyrimidine biosynthesis vitro, using recombinantly expressed enzymes...
Long-range scalar 5J(H1',F) couplings were observed in 5-fluoropyrimidine-substituted RNA. We developed a novel S3E-19F-alpha,beta-edited NOESY experiment for quantitation of these long-range couplings, where the J-couplings can be extracted from inspection intraresidual (H1',H6) NOE cross-peaks. Quantum chemical calculations exploited to investigate relation between and conformations around glycosidic bond oligonucleotides. The theoretical dependence on torsion angle chi described by...
The production of isotopically labeled RNA remains critical to current NMR structural studies. One approach obtain simple spectra is label with a nucleus that not naturally occurring in RNA. Fluorine-19 can serve as sensitive site-specific probe upon incorporation into Here we report the efficient vitro enzymatic synthesis 2-fluoroadenosine-5'-triphosphate and its HIV-2 transactivation region (TAR) by DNA template-directed transcription using phage T7 polymerase. We provide unequivocal...
The fluorescent nucleotide analogue 8-azaguanosine-5'-triphosphate (8azaGTP) is prepared easily by in vitro enzymatic synthesis methods. 8azaGTP an efficient substrate for T7 RNA polymerase and incorporated specifically opposite cytosine the transcription template, as expected a nucleobase with same Watson−Crick hydrogen bonding face guanine. 8-Azaguanine (8azaG) oligonucleotides also recognized guanine during ribonuclease T1 digestion. Moreover, replacement of 8azaG does not alter melting...
Active-site guanines that occupy similar positions have been proposed to serve as general base catalysts in hammerhead, hairpin, and glmS ribozymes, but no specific roles for these demonstrated conclusively. Structural studies place G33(N1) of the ribozyme Bacillus anthracis within hydrogen-bonding distance 2′-OH nucleophile. Apparent pKa values determined from pH dependence cleavage kinetics wild-type mutant ribozymes do not support a role G33, or any other active-site guanine, catalysis....
Understanding how self-cleaving ribozymes mediate catalysis is crucial in light of compelling evidence that human and bacterial gene expression can be regulated through RNA self-cleavage. The hairpin ribozyme catalyzes reversible phosphodiester bond cleavage a mechanism does not require divalent metal cations. Previous structural biochemical implicated the amidine group an active site adenosine, A38, pH-dependent step catalysis. We developed way to determine microscopic pK(a) values based on...
Galectins are glycan-binding proteins translating the sugar-encoded information of cellular glycoconjugates into physiological activities, including immunity, cell migration, and signaling. also interact with non-glycosylated partners in extracellular milieu, among which pre-B receptor (pre-BCR) during B development. How these interactions might interplay glycan-decoding function galectins is unknown. Here, we perform NMR experiments on native membranes to monitor Gal-1 binding surface...
While the number of characterized radical S-adenosyl-l-methionine (SAM) enzymes is increasing, roles these in catalysis remain largely ambiguous. In SAM enzymes, slow initiation step kinetically masks subsequent steps, making it impossible to study kinetics chemistry. Due this kinetic masking, unknown whether reactions require rate acceleration by enzyme active site. Here, we report first evidence that a MoaA accelerates radical-mediated C–C bond formation. catalyzes an unprecedented...
The reactions of indoline (I) and tetrahydroquinoline (THQ) with Os3(CO)10(CH3CN)2 (1) have been studied. Reaction 1 I at ambient temperatures gives Os3(CO)10(μ-H)(μ-η2-C8H7NH) (2), which decarbonylates thermally to give a mixture the tautomeric complexes Os3(CO)9(μ-H)2(μ3-η2-C8H7N) (3 4) whose structures differ by having μ-alkylidene−imino bonding mode (3) vs μ-amido−aryl (4). conversion 2 3 4 follows strictly first-order kinetics equilibrium constant K(4/3) = 6. Further thermolysis or...
Bioassay-guided fractionation of the chloroform/methanol extract a dart poison from Indonesian Borneo (Kalimantan), derived Antiaris toxicaria latex, has led to isolation new cardenolide, toxicarioside A [1]. The structure 1 was deduced by analysis spectroscopic data and first assignment 1H- 13C-NMR shifts for an antiarigenin aglycone. bioassay employed isolate cardenolide involves inhibition Na+/K+-ATPase mimics suspected mode action these "cardiac-glycoside" toxins.
LC/MS analysis of ribonucleosides is traditionally performed by reverse phase chromatography on silica based C18 type stationary phases using MS compatible buffers and methanol or acetonitrile gradients. Due to the hydrophilic polar nature nucleosides, down-scaling analytical methods a two-column nano-flow setup inherently difficult. We present nano-chip ion-trap strategy for routine characterization RNA nucleosides in fmol range. Nucleosides were analyzed positive ion mode 75 μ × 150 mm, 5...
ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTComparative study of the reactions diazomethane with .mu.3-imidoyl and .mu.3-butyne trinuclear clustersMichael Day, William Freeman, Kenneth I. Hardcastle, Mark Isomaki, Shariff E. Kabir, Tim McPhillips, Edward Rosenberg, Lincoln G. Scott, Erich WolfCite this: Organometallics 1992, 11, 10, 3376–3384Publication Date (Print):October 1, 1992Publication History Published online1 May 2002Published inissue 1 October...