16 research outputs found
Solution Structure and Energy Calculation of Bis-Intercalation of Homodimeric Thiazole Orange Dye Derivatives in DNA: Effects of Modifying the Linker
No full textWe have used two-dimensional 1H NMR spectroscopy obtained at 750 MHz to determine a high-resolution solution structure of the double-stranded DNA oligonucleotide d(5‘-CGCTAGCG-3‘)2
complexed with the bis-intercalating dye 1,1‘-(5,5,9,9-tetramethyl-5,9-diazatridecamethylene)-bis-4-[3-ethyl-2,3-dihydro(benzo-1,3-thiazolyl)-2-methylidene]quino-linium tetraiodide (TOTO11Et). The
determination of the structure was based on a complete relaxation matrix analysis of the NOESY
cross-peaks followed by restrained molecular dynamics calculations. Forty final structures were
generated for the TOTO11Et complex from A-form and B-form dsDNA starting structures. The root-mean-square (rms) deviation of the coordinates for the 40 structures of the complex was 0.52 Å. A
conformational analysis of the deoxyribose rings based on coupling constants obtained from selective
DQF-COSY spectra revealed that all ring conformations were almost pure S-type. The structure of
the TOTO11Et complex was compared with the structure of a similar DNA complex with a dye
containing a shorter linker (TOTOEt). Substantial differences were observed between the two
structures because of the difference in the length of the linker. Most prominent was a large difference
in the degree of unwinding of the dsDNA part in the two complexes. Unwinding of 73° and 22° relative
to the free dsDNA was observed for the complexes with TOTOEt and TOTO11Et, respectively. The
AMBER94 force field together with the GB/SA solvation model was used for energy calculations on
both of the two complexes. In the calculations, the complex formation was divided into two steps: (i)
unwinding of the free oligonucleotide and (ii) association of the bis-intercalators to the unwound
oligonucleotide. The complex formation was in favor of TOTO11Et, mainly because the dsDNA is
distorted less in the complex with TOTO11Et than in the complex with TOTOEt
The Global Conformation of the Hammerhead Ribozyme Determined Using Residual Dipolar Couplings<sup>†</sup>
No full textThe global structure of the hammerhead ribozyme was determined in the absence of Mg2+ by
solution NMR experiments. The hammerhead ribozyme motif forms a branched structure consisting of
three helical stems connected to a catalytic core. The 1H-15N and 1H-13C residual dipolar couplings were
measured in a set of differentially 15N/13C-labeled ribozymes complexed with an unlabeled noncleavable
substrate. The residual dipolar couplings provide orientation information on both the local and the global
structure of the molecule. Analysis of the residual dipolar couplings demonstrated that the local structure
of the three helical stems in solution is well modeled by an A-form conformation. However, the global
structure of the hammerhead in solution in the absence of Mg2+ is not consistent with the Y-shaped
conformation observed in crystal structures of the hammerhead. The residual dipolar couplings for the
helical stems were combined with standard NOE and J coupling constant NMR data from the catalytic
core. The NOE data show formation of sheared G−A base pairs in domain 2. These NMR data were used
to determine the global orientation of the three helical stems in the hammerhead. The hammerhead forms
a rather extended structure under these conditions with a large angle between stems I and II (∼153°), a
smaller angle between stems II and III (∼100°), and the smallest angle between stems I and III (∼77°).
The residual dipolar coupling data also contain information on the dynamics of the molecule and were
used here to provide qualitative information on the flexibility of the helical domains in the hammerhead
ribozyme−substrate complex
Locked Nucleic Acid (LNA) Recognition of RNA: NMR Solution Structures of LNA:RNA Hybrids
No full textLocked nucleic acids (LNAs) containing one or more 2‘-O,4‘-C-methylene-linked bicyclic
ribonucleoside monomers possess a number of the prerequisites of an effective antisense oligonucleotide,
e.g. unprecedented helical thermostability when hybridized with cognate RNA and DNA. To acquire a detailed
understanding of the structural features of LNA giving rise to its remarkable properties, we have conducted
structural studies by use of NMR spectroscopy and now report high-resolution structures of two LNA:RNA
hybrids, the LNA strands being d(5‘-CTGATLATGC-3‘) and d(5‘-CTLGATLATLGC-3‘), respectively, TL
denoting a modified LNA monomer with a thymine base, along with the unmodified DNA:RNA hybrid. In
the structures, the LNA nucleotides are positioned as to partake in base stacking and Watson−Crick base
pairing, and with the inclusion of LNA nucleotides, we observe a progressive change in duplex geometry
toward an A-like duplex structure. As such, with the inclusion of three LNA nucleotides, the hybrid adopts
an almost canonical A-type duplex geometry, and thus it appears that the number of modifications has
reached a saturation level with respect to structural changes, and that further incorporations would furnish
only minute changes in the duplex structure. We attempt to rationalize the conformational steering induced
by the LNA nucleotides by suggesting that the change in electronic density at the brim of the minor groove,
introduced by the LNA modification, is causing an alteration of the pseudorotational profile of the 3‘-flanking
nucleotide, thus shifting this sugar equilibrium toward N-type conformation
Comparison of the ability of wild type and stabilized human IgG4 to undergo Fab arm exchange with endogenous IgG4 in vitro and in vivo
No full textStructural studies of LNA:RNA duplexes by NMR: Conformations and implications for RNase H activity
No full textKem
CS16-3. The extracellular domains of the IL-21 receptor are bridged by glycosylation through conserved anchor points.
No full textRecommended from our members
Unmasking prurigo nodularis endotypes: Dominant role of IL 31 dysregulation
No full textThe existence of multiple conformers of interleukin-21 directs engineering of a superpotent analogue
No full text
