摘要:

A triple helix can be formed upon binding of a pyrimidine oligonucleotide to the major groove of a homopurine-homopyrimidine (R·Y) double-stranded DNA target site. Here, we report that this reaction can be influenced by base methylation. The pyrimidine strand 5′- TmCTmCTmCTmCTTmCT (mY12), whose cytosine residues are methylated at C5, does not bind the duplex 5′-AGAGAGAGAAGA·3′-TCTCTCTCTTCT (R12·Y12) to yield a 12-triad triplex, as would be expected from these DNA sequences. Rather, a complex of overlapping oligonucleotides, which we define concatenamer, is formed. The concatenamer is clearly evidenced by Polyacrylamide gel electrophoresis (PAGE) since it migrates with a smeared band of very low mobility. The stoichiometry of the concatenamer, determined by both UV mixing curves and electrophoresis, is surprisingly found to be (R12· 2mY12)n, thus showing that the unmethylated Y12strand is excluded from the complex. Denaturation experiments performed by ultraviolet absorbance (UV) and differential scanning calorimetry (DSC) show that the concatenamers melt with a single and highly cooperative transition whoseTmstrongly depends on pH. Overall, the data point to the conclusion that the concatenamers are in triple helix, where the methylated mY12strand is engaged in both Watson-Crick and Hoogsteen base pairings, thus displacing the Y12strand from the R12·Y12duplex. A possible mechanism of concatenamer formation is proposed. The results presented in this paper show that 5-methylcytosine brings about a strong stabilizing effect on both double and triple DNA helices, and that pyrimidine oligonucleotides containing 5-methylcytosine can displace from R·Y duplexes the analogous non-methylated strand. The advantage of using methylated oligonucleotides in antisense technology is discussed.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508027

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

Relative humidity induces the reversible crystal transition of guanosine between the dihydrate and the anhydrous state. The characteristics of the transition was investigated by means of X-ray powder diffraction analysis and high-resolution solid-state13C NMR spectroscopy. Adsorption-desorption hysteresis was observed. Guanosine dihydrate (the H-state) which is crystallized from an aqueous solution rapidly loses crystal water below 10% relative humidity (rh), and is anhydrous at 0% rh (the A-state). The crystals gradually recover the H-state at approximate 20% rh. In the adsorption process between 10–20% rh, there exists one intermediate state, M, with 1.2−1.3 moles water per mole guanosine. The lattice of the M-state was determined to be orthorhombic with the cell parameters of a = 16.248(1), b = 11.603(1), and c = 13.643 (2) Å.The base-stacking structure is retained throughout the transition. On the other hand, conformational changes of the riboses and break of the hydrogen-bonding network between the bases would be induced in the A-state in conformity with lack of crystal water.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508028

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

We probed conformational polymorphism of a synthetic DNA poly(dA-dT) by introducing various small amounts of bulky spherical hydrophobic isopropyl groups into the polynucleotide primary structure. For this purpose, three mixed copolymers of poly(dA-dT, ip5dU) were synthesized in which 2.6 %, 8.6 % or 14.2 % of the polynucleotide pyrimidine bases had the isopropyl group in position 5. The isopropyls made the formation of both A-form and X-form incomplete, and this effect increased with the increasing isopropyl amount in the polynucleotide. However, the polynucleotide isomerization into the A-form was hindered by the isopropyls while the isomerization into the X-form was rather promoted. This observation indicates that, unlike the A-form, the X-form has the base pairs shifted towards the double helix major groove. Z-form was also promoted by the lowest concentration of the isopropyl groups while the most isopropylated poly(dA-dT) aggregated under the Z-form inducing conditions.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508029

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

X-ray diffraction data from well oriented and polycrystalline fibers of the lithium salt of poly d(AATT)·poly d(AATT) are isomorphous with those from B-DNA The double-helix consists of conformationally identical antiparallel strands and the molecular symmetry is 2 52; the asymmetric unit is a tetranucleotide, AATT, and 5 tetranucleotides span two turns per strand. Two double helices pass through a monoclinic unit cell of dimensions a = 31.05, b = 22.62, c = 33.85 Å (fiber axis) and γ= 90°. In each repeating motif, the four nucleotides have distinct conformations, TpA displays an axial P-O bond and there is shortening of minor groove in the central region.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508030

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

A novel method mapping the DNA or RNA sequence into a folding curve in three dimensional space, the Z curve, has been proposed based on the symmetry of the regular tetrahedrons. There exists a unique Z curve for a given DNA sequence, on the contrary, the DNA sequence can be uniquely determined by the given Z curve. The properties of the Z curves have been studied in great details. The symmetry, the periodicity, the local motif, and the global feather of the distribution of bases of the DNA sequences are reflected by the rich folding structures of the Z curves. The Z curves may be smoothed by the B-spline functions of different orders. Therefore, the Z curves may have any resolution by choosing the suitable spline functions. The higher the order of the B-spline function chosen, the lower the resolution of the Z curve. So, the Z curves are suitable for visualizing and analyzing the DNA sequences with any length. The study of the Z curves develops further a new area to visualizing and analyzing the DNA sequences by a geometrical approach. The method of the Z curves may be strengthened by using the ripe mathematical tools of geometry on the one hand; and by using the powerful technique of the computer graphics on the other hand.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508031

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

The method of DNA sequencing by hybridization with oligonucleotides matrix (SHOM) developed in this laboratory (1,2) uses the matrix of oligonucleotides immobilized within Polyacrylamide gel. The particular feature of this matrix is that the apparent thermostability of the duplexes depends on the concentration of gel-immobilized oligonucleotides. This dependence is specific for oligonucleotides immobilized in the gel volume (3-D-immobilization) rather than on a flat surface of a filter or glass (2-D-immobilization). The theory has been developed that provides a quantitative description of temperature-dependent duplex dissociation within gel. The theory takes into account that the diffusion of dissociated DNA out of the gel is retarded by multiple acts of association-dissociation of DNA with immobilized oligonucleotides. It allows to calculate the apparent dissociation temperature of duplexes and describes quantitatively its growth upon increase in the enthalpy of duplex dissociation, concentration of immobilized oligonucleotides, gel thickness and decrease of dissociation entropy and washing time. Concentration of gel-immobilized oligonucleotides can be calculated for a normalized matrix in which GC-rich and AT-rich duplexes exhibit the same apparent thermostabilities and are washed off at the same temperature. This simplifies identification of perfect duplexes formed on the matrix which can be carried out for all duplexes at the same temperature. The gel-immobilized oligonucleotide matrix provides also a higher capacity for immobilization and therefore a higher sensitivity of measurements, resulting in a higher discrimination power for identification of perfect duplexes as compared with matrixes of oligonucleotides immobilized on a surface.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508032

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

In this paper we consider the efficiency of additional rounds of “continuous stacking” hybridization in DNA sequence reconstruction by hybridization with oligonucleotide matrix (SHOM). After the initial hybridization of target DNA with the matrix of oligonucleotides of fixed lengthLsome additional hybridizations should be carried out in the presence of fluorescently labeled oligonucleotides of another length l. These additional oligonucleotides can hybridize in tandem with matrix tuples (continuous stacking hybridization) thus forming an extended duplex with the target DNA strand. The additional data obtained allows resolutions of branching points arising in the reconstruction procedure. Multiple rounds of continuous stacking hybridization considerably increase the efficiency of the sequencing method, eventually approaching the power of (L+l)-matrix. We develop here an algorithm that allows us to minimize the number of additional hybridization steps, by assembling sets of l-tuples to be added together in each round of continuous stacking hybridization. For SHOM using a matrix of octanucleotides, continuous stacking hybridization with pen- tanucleotides increases the length of unambiguously sequenced DNA from 200 to several thousands of base pairs.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508033

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

Our analysis of the data of van den Engh, Sachs, and Trask(Science 257, 1410 (1992)), for the dependence of the mean square distance between pairs of hybridization sites (<Ln2>,μm2)on the known genomic distance (n, bp) separating these sites on chromosome number 4 in G1 human fibroblast nuclei, shows that <Ln2> is proportional to n2vwithv= 3/5 for n < 1 Mbp. Thev-value of 3/5 is characteristic of flexible polymer chains with excluded volume effects in dilute good solutions. Since the DNA concentration in nuclei is very high (ca.1–10 mg/m1), and theory (Flory,J. Chem. Phys. 17, 303, 1949) predictsv= 1/2 for overlapping polymers, the finding ofv= 3/5 means that the chromatin fibers do not overlap in interphase nuclei. The dependence of <Ln2> on n for n < 4 Mbp is consistent with the model of large (∼ 6 Mbp, 3μm diameter) loops of interphase chromatin attached to nuclear membrane sites. Using the constant (e.g., Widom,Ann. Rev. Biophys. Biophys. Chem. 18, 365 (1989)) and variable (Williams & Langmore,Biophys. J. 59, 606 (1991)) diameter fiber models, the Kuhn statistical segment of the 30 nm chromatin fiber was estimated to have a length of 196–272 nm with a corresponding DNA content of 21–37 kbp. Based on the model of Shimada and Yamakawa (Macromolecules 17, 689 (1984);Biopolymers 27, 651 (1988)) for circular wormlike chains, we estimated the most favorable size of the small loops of the 30 nm fiber to be 36–62 kbp with a diameter of 94–131 nm. Both the size and diameter estimates are consistent with experimental measurements from the literature: 60 kbp for average loop size (van Holde,Chromatin, Ch. 7, Springer-Verlag, New York, 1989) and 125 nm for the diameter (Belmontet al., Chromosome 98, 129 (1989)).

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508034

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

The 60 amino acid proline-rich neutralization domain of the external surface unit glycoprotein of feline leukemia virus was chemically synthesized in total and in fragments. We examined the ability of these retroviral peptides to form ordered conformations using1H- NMR, circular dichroism spectroscopy, and intrinsic viscosity measurements. One dimensional nuclear magnetic resonance spectroscopy revealed that the 60 amino acid peptide could form a stable, folded structure that was long-lived, as shown by the ability to protect amide-protons in D20. Peptides corresponding to the N-terminal 42, N-terminal 20 amino acids, and middle 20 amino acid sections could also form stable structures. The C-terminal segment did not protect any protons in D20. Interestingly, self assembly of the N-terminal 42 and C-terminal 16 amino acid peptides into a structure very close to that of the 60 amino acid domain was observed. The circular dichroism results reveals a large negative cotton effect at 198 nm that is characteristic of the proline-rich β-turn helixes which consist predominantly oftrans-proline. The intrinsic viscosity results suggest a non-random coil structure that is rod shaped. Our conclusion is that PRN60 forms a β-turn helix and that this region of FeLV- gp70 is a separate folding domain of the retroviral surface unit glycoprotein. The unique conformational properties of PRN60 and its critical role as the predominant target for neutralizing antibody responses suggest that this peptide is a reasonable candidate for producing a synthetic peptide vaccine for FeLV.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508035

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor

摘要:

Coconut foliar decay virus (CFDV) contains the single-stranded circular DNA molecules of 1291 nucleotides which were found to replicate autonomously in the cells of the diseased palms. The special features of the CFDV DNA sequence, including putative secondary structure and the distribution of the inverted repeat motifs, are investigated with computer-assisted prediction methods. It is evident that the structural principle of the branched series of long and short double helixes interspersed by short non-helical regions is existed for CFDV virion DNA. The total degree of base pairing is near 62%. We have also predicted the presence of several sequence elements formed by inverted repeat motifs which are potentially capable of binding the eukaryotic transcriptional regulatory factors.

ISSN:0739-1102    

DOI:10.1080/07391102.1994.10508036

出版商:Taylor & Francis Group    

年代:1994

数据来源: Taylor