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1.
We have determined the stability of intramolecular DNA quadruplexes in which the four G3-tracts are connected by non-nucleosidic linkers containing propanediol, octanediol or hexaethylene glycol, replacing the TTA loops in the human telomeric repeat sequence. We find that these sequences all fold to form intramolecular complexes, which are stabilized by lithium < sodium < potassium. Quadruplex stability increases in the order propanediol < hexaethylene glycol < octanediol. The shallower shape of the melting profile with propanediol linkers and its lower dependency on potassium concentration suggests that this complex contains fewer stacks of G-quartets. The sequence with octanediol linkers displays a biphasic melting profile, suggesting that it can adopt more than one stable structure. All these complexes display melting temperatures above 310 K in the presence of 10 mM lithium, without added potassium, in contrast to the telomeric repeat sequence. These complexes also fold much faster than the telomeric repeat and there is little or no hysteresis between their melting and annealing profiles. In contrast, the human telomeric repeat sequence and a complex containing two hexaethylene glycol groups in each loop, are less stable and fold more slowly. The melting and annealing profiles for the latter sequence show significant differences, even when heated at 0.2°C min–1. CD spectra for the oligonucleotides containing non-nucleosidic linkers show positive maxima at 264 nm, with negative minima ~244 nm, which are characteristic of parallel quadruplex structures. These results show that the structure and stability of intramolecular quadruplexes is profoundly influenced by the length and composition of the loops. 相似文献
2.
Hammerhead ribozymes were synthesized in which the tetranucleotide loop II was replaced by non-nucleotidic linkers of 7, 13, 17 and 19 atoms length. Ribozymes with 17 and 19 atom linkers, in combination with a 4 base pair stem II, had catalytic efficiencies which were 2 fold increased to that of the parent ribozyme with a tetranucleotide loop. Ribozymes with these linkers, but in combination with a 2 base pair stem II, showed a 2 fold decrease in catalytic efficiency when compared to the parent ribozyme. Prolonged preincubation in the presence of MgCl2 was required for hexaethylene glycol linker-modified ribozymes to obtain maximum activity and reproducible kinetic data. 相似文献
3.
We have examined the properties of intramolecular G-quadruplexes in which the G3 tracts are separated by single base loops. The most stable complex contained 1',2'-dideoxyribose in all three loops, while loops containing T and C were slightly less stable (by about 2 degrees C). Quadruplexes containing loops with single A residues were less stable by 8 degrees C for each T to A substitution. These folded sequences display similar CD spectra, which are consistent with the formation of parallel stranded complexes with double-chain reversal loops. These results demonstrate that loop sequence, and not just length, affects quadruplex stability. 相似文献
4.
Circular dichroism studies of an oligodeoxyribonucleotide containing a hairpin loop made of a hexaethylene glycol chain: conformation and stability. 总被引:13,自引:11,他引:2
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M Durand K Chevrie M Chassignol N T Thuong J C Maurizot 《Nucleic acids research》1990,18(21):6353-6359
An oligodeoxyribonucleotide, d(GCTCACAAT-X-ATTGTGAGC), where X represents a hexaethylene glycol chain, was studied using circular dichroism spectroscopy. Its conformation and conformational stability were compared to those of compounds where X was replaced by four thymines and to the duplex of same sequence without loop. The compound with the hexaethylene glycol chain can form a hairpin looped structure as well as a bulged duplex structure. In both cases the duplex region of the oligodeoxyribonucleotide exhibits the same conformation. In similar conditions the oligodeoxyribonucleotide with a four thymines loop forms exclusively a hairpin structure. Comparison between the thermodynamic parameters (delta H, delta S, delta G) associated with the formation of the structure of the three compounds are presented. In the case of the compound with the hexaethylene glycol chain it is shown that the large increase in its melting temperature (by about 35 degrees C in our experimental conditions) when compared to the non looped structure is mainly due to the fact that its melting process is intramolecular (monomolecular) whereas the other one is bimolecular. 相似文献
5.
Sun H Tang Y Xiang J Xu G Zhang Y Zhang H Xu L 《Bioorganic & medicinal chemistry letters》2006,16(13):3586-3589
Quercetin is a kind of flavonoid which has been proved to exhibit anti-tumor activity. The interaction modes of quercetins with monomeric and dimeric G-quadruplexes were studied by absorption, fluorescence, CD, and (1)H NMR spectroscopies. The ligands were found to be stacked with terminal tetrads of monomeric G-quadruplexes by intercalation and bound to dimeric G-quadruplexes by groove binding. 相似文献
6.
G-rich oligonucleotides T30695 (or T30923), with the sequence of (GGGT)(4), and T40214, with the sequence of (GGGC)(4), have been reported to exhibit anti-HIV and anticancer activity. Here we report on the structure of a dimeric G-quadruplex adopted by a derivative of these sequences in K(+) solution. It comprises two identical propeller-type parallel-stranded G-quadruplex subunits each containing three G-tetrad layers that are stacked via the 5'-5' interface. We demonstrated control over the stacking of the two monomeric subunits by sequence modifications. Our analysis of possible structures at the stacking interface provides a general principle for stacking of G-quadruplexes, which could have implications for the assembly and recognition of higher-order G-quadruplex structures. 相似文献
7.
Biochemical and biophysical studies on the folding of the core region of the origin of replication of bacteriophage M13.
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A van Belkum M J Blommers H van den Elst J H van Boom C W Hilbers 《Nucleic acids research》1990,18(16):4703-4710
DNA oligonucleotides with the sequence corresponding to the plus strand origin of replication of the filamentous bacteriophage M13 are studied. Biochemical structure probing and UV melting studies, supplemented with initial NMR experiments, are used to investigate structural features of a 51-nucleotides long synthetic oligonucleotide and two oligonucleotides that are integral parts of this latter molecule. The results demonstrate the feasibility and complementarity of the use of methidiumpropyl.EDTA-Fe(II) and nuclease S1 in the structural analysis of small oligonucleotides. The bacteriophage origin region appears to comprise two hairpins. The first hairpin, which contains a cleavage site for the bacteriophage gene II protein, has a large and probably flexible loop. NMR as well as UV melting studies demonstrate that the second hairpin contains a stable three-membered loop. Both hairpins are present in the 51-mer, which forms a stable tertiary structure. 相似文献
8.
It is shown that component analysis could be applied to study the UV difference spectra of cytidine oligomers and hairpin oligonucleotides with cytidines in the loop region in order to account for the melting and titration results in terms of cytidine stacking and protonation. Upon acid titration, the dC(10) oligomer undergoes cooperative conformational transition at pH 6.3 accompanied by protonation and formation of the i-structure with half of the residues protonated. The stability of the hemiprotonated structure increases with decreasing pH, the i-structure persisting still in the region of pH相似文献
9.
Ahmed A. El-Sayed Erik B. Pedersen Nahid A. Khaireldin 《Nucleosides, nucleotides & nucleic acids》2013,32(12):872-879
Certain cytosine-rich (C-rich) DNA sequences can fold into secondary structures as four-stranded i-motifs with hemiprotonated base pairs. Here we synthesized C-rich TINA-intercalating oligonucleotides by inserting a nonnucleotide pyrene moiety between two C-rich regions. The stability of their i-motif structures was studied by using UV melting temperature measurements and circular dichroism spectra at different pH values under noncrowding and crowding conditions (20% poly(ethylene glycol)). When TINA ((R)-3-((4-(1-pyrenylethynyl)benzyl)oxy) propane-1,2-diol) is inserted, the oligonucleotides could form an i-motif at a higher pH than observed for the corresponding wildtype oligonucleotide. 相似文献
10.
Multinuclear NMR studies of DNA hairpins. 1. Structure and dynamics of d(CGCGTTGTTCGCG) 总被引:9,自引:0,他引:9
The solution structure of the hairpin formed by d(CGCGTTGTTCGCG) has been examined in detail by a wide variety of NMR techniques. The hairpin was characterized by proton NMR to obtain interproton distances and torsion angle information. An energy-minimized model was constructed that is consistent with these data. The hairpin consists of a B-DNA stem of four C-G base pairs and a loop region consisting of five unpaired bases. Three bases in the 5' of the loop are stacked over the 3' end of the stem, and the other two bases in the 3' of the loop are stacked over the 5' end of the stem. The phosphorus NMR spectrum revealed a phosphate in the stem region with an unusual conformation, and two phosphates, P9 and P10, were found to undergo intermediate exchange between conformations. The hairpin was also synthesized with a carbon-13 label in each of the thymidine C6 carbons, and relaxation measurements were performed to determine the extent of internal motions in the loop region. The loop bases are more flexible than the stem bases and exhibit subnanosecond motions with an amplitude corresponding to diffusion in a cone of approximately 30 degrees. 相似文献
11.
A sequence-independent study of the influence of short loop lengths on the stability and topology of intramolecular DNA G-quadruplexes 总被引:3,自引:0,他引:3
G-Rich sequences found within biologically important regions of the genome have been shown to form intramolecular G-quadruplexes with varied loop lengths and sequences. Many of these quadruplexes will be distinguishable from each other on the basis of their thermodynamic stabilities and folded conformations. It has been proposed that loop lengths can strongly influence the topology and stability of intramolecular G-quadruplexes. Previous studies have been limited to the analysis of quadruplex sequences with particular loop sequences, making it difficult to make generalizations. Here, we describe an original study that aimed to elucidate the effect of loop length on the biophysical properties of G-quadruplexes in a sequence-independent context. We employed UV melting and circular dichroism spectroscopy to examine and compare the properties of 21 DNA quadruplex libraries, each comprising partially randomized loop sequences with lengths ranging from one to three nucleotides. Our work supports a number of general predictions that can be made solely on the basis of loop lengths. In particular, the results emphasize the strong influence of single-nucleotide loops on quadruplex properties. This study provides a predictive framework that may help identify or classify biologically relevant G-quadruplex-forming sequences. 相似文献
12.
Solution NMR and computer simulation studies of active site loop motion in triosephosphate isomerase
Solution NMR spin relaxation experiments and classical MD simulations are used to study the dynamics of triosephosphate isomerase (TIM) in complex with glycerol 3-phosphate (G3P). Three regions in TIM exhibit conformational transitions on the micros-ms time scale as detected by chemical exchange broadening effects in NMR spectroscopy: residue Lys 84 on helix C, located at the dimeric interface; active site loop 6; and helix G. The results indicate that the conformational exchange process affecting the residues of loop 6 is the correlated opening and closing of the loop. Distinct processes are responsible for the chemical exchange linebroadening observed in the other regions of TIM. MD simulations confirm that motions of individual residues within the active site loop are correlated and suggest that the chemical exchange processes observed for residues in helix G arise from transitions between 3(10)- and alpha-helical structures. The results of the joint NMR and MD study provide global insight into the role of conformational dynamic processes in the function of TIM. 相似文献
13.
We studied the structures and stabilities of G-quadruplexes formed in Myc1234, the region containing the four consecutive 5' runs of guanines of c-MYC promoter NHE III(1,) which have recently been shown to form in a supercoiled plasmid system in aqueous solution. We determined the NMR solution structure of the 1:2:1 parallel-stranded loop isomer, one of the two major loop isomers formed in Myc1234 in K(+) solution. This major loop isomer, although sharing the same folding structure, appears to be markedly less stable than the major loop isomer formed in the single-stranded c-MYC NHE III(1) oligonucleotide, the Myc2345 G-quadruplex. Our NMR structures indicated that the different thermostabilities of the two 1:2:1 parallel c-MYC G-quadruplexes are likely caused by the different base conformations of the single nucleotide loops. The observation of the formation of the Myc1234 G-quadruplex in the supercoiled plasmid thus points to the potential role of supercoiling in the G-quadruplex formation in promoter sequences. We also performed a systematic thermodynamic analysis of modified c-MYC NHE III(1) sequences, which provided quantitative measure of the contributions of various loop sequences to the thermostabilities of parallel-stranded G-quadruplexes. This information is important for understanding the equilibrium of promoter G-quadruplex loop isomers and for their drug targeting. 相似文献
14.
NMR and CD data have previously shown the formation of the T(4) tetraloop hairpin in aqueous solutions, as well as the possibility of the B-to-Z transition in its stem in high salt concentration conditions. It has been shown that the stem B-to-Z transition in T(4) hairpins leads to S (south)- to N (north)-type conformational changes in the loop sugars, as well as anti to syn orientations in the loop bases. In this article, we have compared by means of UV absorption, CD, Raman, and Fourier transform infrared (FTIR), the thermodynamic and structural properties of the T(4) and A(4) tetraloop hairpins formed in 5'-d(CGCGCG-TTTT-CGCGCG)-3' and 5'-d(CGCGCG-AAAA-CGCGCG)-3', respectively. In presence of 5M NaClO(4), a complete B-to-Z transition of the stems is first proved by CD spectra. UV melting profiles are consistent with a higher thermal stability of the T(4) hairpin compared to the A(4) hairpin. Order-to-disorder transition of both hairpins has also been analyzed by means of Raman spectra recorded as a function of temperature. A clear Z-to-B transition of the stem has been confirmed in the T(4) hairpin, and not in the A(4) hairpin. With a right-handed stem, Raman and FTIR spectra have confirmed the C2'-endo/anti conformation for all the T(4) loop nucleosides. With a left-handed stem, a part of the T(4) loop sugars adopt a N-type (C3'-endo) conformation, and the C3'-endo/syn conformation seems to be the preferred one for the dA residues involved in the A(4) tetraloop. 相似文献
15.
M. J. Bennett S. Choe D. Eisenberg 《Protein science : a publication of the Protein Society》1994,3(9):1444-1463
The refined structure of dimeric diphtheria toxin (DT) at 2.0 A resolution, based on 37,727 unique reflections (F > 1 sigma (F)), yields a final R factor of 19.5% with a model obeying standard geometry. The refined model consists of 523 amino acid residues, 1 molecule of the bound dinucleotide inhibitor adenylyl 3'-5' uridine 3' monophosphate (ApUp), and 405 well-ordered water molecules. The 2.0-A refined model reveals that the binding motif for ApUp includes residues in the catalytic and receptor-binding domains and is different from the Rossmann dinucleotide-binding fold. ApUp is bound in part by a long loop (residues 34-52) that crosses the active site. Several residues in the active site were previously identified as NAD-binding residues. Glu 148, previously identified as playing a catalytic role in ADP-ribosylation of elongation factor 2 by DT, is about 5 A from uracil in ApUp. The trigger for insertion of the transmembrane domain of DT into the endosomal membrane at low pH may involve 3 intradomain and 4 interdomain salt bridges that will be weakened at low pH by protonation of their acidic residues. The refined model also reveals that each molecule in dimeric DT has an "open" structure unlike most globular proteins, which we call an open monomer. Two open monomers interact by "domain swapping" to form a compact, globular dimeric DT structure. The possibility that the open monomer resembles a membrane insertion intermediate is discussed. 相似文献
16.
A prolonged expansion of GGGGCC repeat within non-coding region of C9orf72 gene has been identified as the most common cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), which are devastating neurodegenerative disorders. Formation of unusual secondary structures within expanded GGGGCC repeat, including DNA and RNA G-quadruplexes and R-loops was proposed to drive ALS and FTD pathogenesis. Initial NMR investigation on DNA oligonucleotides with four repeat units as the shortest model with the ability to form an unimolecular G-quadruplex indicated their folding into multiple G-quadruplex structures in the presence of K+ ions. Single dG to 8Br-dG substitution at position 21 in oligonucleotide d[(G4C2)3G4] and careful optimization of folding conditions enabled formation of mostly a single G-quadruplex species, which enabled determination of a high-resolution structure with NMR. G-quadruplex structure adopted by d[(G4C2)3GGBrGG] is composed of four G-quartets, which are connected by three edgewise C-C loops. All four strands adopt antiparallel orientation to one another and have alternating syn-anti progression of glycosidic conformation of guanine residues. One of the cytosines in every loop is stacked upon the G-quartet contributing to a very compact and stable structure. 相似文献
17.
A DNA hairpin with a single residue loop closed by a strongly distorted Watson-Crick G x C base-pair
El Amri C Mauffret O Monnot M Tevanian G Lescot E Porumb H Fermandjian S 《Journal of molecular biology》1999,294(2):427-442
Our previous NMR and modeling studies have shown that the single-stranded 19mer oligonucleotides d(AGCTTATC-ATC-GATAA GCT) -ATC- and d(AGCTTATC-GAT-GATAAGCT) -GAT- encompassing the strongest topoisomerase II cleavage site in pBR322 DNA could form stable hairpin structures. A new sheared base-pair, the pyrimidine-purine C x A, was found to close the single base -ATC- loop, while -GAT- displayed a flexible loop of three/five residues with no stabilizing interactions. Now we report a structural study on -GAC-, an analog of -GAT-, derived through the substitution of the loop residue T by C. The results obtained from NMR, non-denaturing PAGE, UV-melting, circular dichroism experiments and restrained molecular dynamics indicate that -GAC- adopts a hairpin structure folded through a single residue loop. In the -GAC- hairpin the direction of the G9 sugar is reversed relative to the C8 sugar, thus pushing the backbone of the loop into the major groove. The G9 x C11 base-pair closing the loop is thus neither a sheared base-pair nor a regular Watson-Crick one. Although G9 and C11 are paired through hydrogen bonds of Watson-Crick type, the base-pair is not planar but rather adopts a wedge-shaped geometry with the two bases stacked on top of each other in the minor groove. The distortion decreases the sugar C1'-C1' distance between the paired G9 and C11, to 8 A versus 11 A in the standard B-DNA. The A10 residue at the center of the loop interacts with the G9 x C11 base-pair, and seems to contribute to the extra thermal stability displayed by -GAC- compared to -GAT-. Test calculations allowed us to identify the experimental NOEs critical for inducing the distorted G.C Watson-Crick base-pair. The preference of -GAC- for a hairpin structure rather than a duplex is confirmed by the diffusion constant values obtained from pulse-field gradient NMR experiments. All together, the results illustrate the high degree of plasticity of single-stranded DNAs which can accommodate a variety of turn-loops to fold up on themselves. 相似文献
18.
Influence of dangling thymidine residues on the stability and structure of two DNA duplexes 总被引:5,自引:0,他引:5
We have employed temperature-dependent UV spectroscopy, circular dichroism (CD), 400-MHz proton nuclear magnetic resonance (NMR), and computer modeling to characterize both structurally and thermodynamically the influence of unpaired, dangling thymidine residues (T) on the thermal stability and melting behavior of two DNA core duplexes. The specific DNA double helices that we have investigated in this work are core duplexes [d(GC)3]2 (I) and [d(CG)3]2 (IV), 3' dangling T derivatives [d[(GC)3TT]]2 (II) and [d[(CG)3TT]]2 (V), and 5' dangling T derivatives [d[TT(GC)3]]2 (III) and [d[TT(CG)3]]2 (VI). Our experimental data allow us to reach the following conclusions: (1) For both core duplexes (I and IV), the addition of dangling T residues on either the 5' or 3' end causes an increase in the optical melting temperature tm. (2) For both core duplexes, 5' dangling T residues induce a greater increase in the optical tm's than 3' dangling T residues. (3) For both cores duplexes, the increase in tm induced by the addition of dangling T residues is enthalpic in origin, with 5' dangling T residues inducing a greater increase in the van't Hoff transition enthalpy than 3' dangling T's. (4) Dangling T residues cause downfield shifts in all of the nonexchangeable aromatic protons of the [d(GC)3]2 core duplex (I), with the 5' T residues inducing the largest shifts. For the most part, this trend does not hold with the [d(CG)3]2 core duplex (IV). (5) For both core duplexes, the addition of dangling T residues causes an increase in the NMR tm's of almost all the nonexchangeable aromatic protons of the core duplex.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
19.
Jielin Chen Mingpan Cheng Gilmar
F Salgado Petr Stadlbauer Xiaobo Zhang Samir Amrane Aurore Gudin Fangni He Jií poner Huangxian Ju Jean-Louis Mergny Jun Zhou 《Nucleic acids research》2021,49(16):9548
Genomic sequences susceptible to form G-quadruplexes (G4s) are always flanked by other nucleotides, but G4 formation in vitro is generally studied with short synthetic DNA or RNA oligonucleotides, for which bases adjacent to the G4 core are often omitted. Herein, we systematically studied the effects of flanking nucleotides on structural polymorphism of 371 different oligodeoxynucleotides that adopt intramolecular G4 structures. We found out that the addition of nucleotides favors the formation of a parallel fold, defined as the ‘flanking effect’ in this work. This ‘flanking effect’ was more pronounced when nucleotides were added at the 5′-end, and depended on loop arrangement. NMR experiments and molecular dynamics simulations revealed that flanking sequences at the 5′-end abolish a strong syn-specific hydrogen bond commonly found in non-parallel conformations, thus favoring a parallel topology. These analyses pave a new way for more accurate prediction of DNA G4 folding in a physiological context. 相似文献
20.
G-Quadruplexes are noncanonical nucleic acid secondary structures based on guanine association that are readily adopted by G-rich RNA and DNA sequences. Naturally occurring genomic G-quadruplex-forming sequences have functional roles in biology that are mediated through structure. To appreciate how this is achieved, an understanding of the likelihood of G-quadruplex formation and the structural features of the folded species under a defined set of conditions is informative. We previously systematically investigated the thermodynamic stability and folding topology of DNA G-quadruplexes and found a strong dependence of these properties on loop length and loop arrangement [Bugaut, A., and Balasubramanian, S. (2008) Biochemistry 47, 689-697]. Here we report on a complementary analysis of RNA G-quadruplexes using UV melting and circular dichroism spectroscopy that also serves as a comparison to the equivalent DNA G-quadruplex-forming sequences. We found that the thermodynamic stability of G-quadruplex RNA can be modulated by loop length while the overall structure is largely unaffected. The systematic design of our study also revealed subtle loop length dependencies in RNA G-quadruplex structure. 相似文献