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1.
Poly(C,A) random copolymer templates direct the oligomerization of 2-MeImpG (2-MeImpX is the 5'-phospho-2-methylimidazolide of the nucleoside X) and 2-MeImpU, resulting in the production of a variety of oligo (G,U)s. This reaction is less efficient than comparable reactions involving poly(C,U) or poly(C,G) templates. The efficiency of monomer incorporation into newly synthesized oligomers is lower for 2-MeImpU than 2-MeImpG, and cannot be improved by increasing the concentration of 2-MeImpU relative to 2-MeImpG. This suggests that RNA templates containing runs of consecutive adenine residues would not be suitable for use in a chemical self-replicating system. The distribution of oligomeric products can be characterized in detail using high-pressure liquid chromatography on an RPC-5 column. Oligomers are separated on the basis of chain length, base composition, and phosphodiester-linkage isomerism. Oligomers up to about the 13-mer, with base composition Gn, Gn-1, U, and Gn-2, U2, have been identified.  相似文献   

2.
Poly(C, G) random copolymer templates direct the oligomerization of 2-Me-ImpG and 2-MeImpC, resulting in the production of a variety of oligo(G, C)s. The efficiency of monomer incorporation into newly synthesized oligomers is greater for 2-MeImpG than for 2-MeImpC, and decreases for both monomers as the guanine content of the template increases. The relatively low efficiency of oligomerization on guanine-rich templates is largely a consequence of intra- and intermolecular template self-structure. The problem of template self-structure is clearly a major obstacle to the development of a system of self-replicating polynucleotides. The distribution of oligomeric products can be characterized in detail using high-pressure liquid chromatography on an RPC-5 column. Oligomers are separated on the basis of chain length, base composition and phosphodiester-linkage isomerism. Oligomers up to about the 12-mer, with base composition Gn, Gn-1C and Gn-2C2, have been identified. The 3' to 5' regiospecificity of the products is high, particularly for oligomers with base composition Gn.  相似文献   

3.
Summary A random copolymer, poly(CA), containing approximately equal amounts of cytidine (C) and adenosine (A), when incubated with a mixture of guanosine-5-phosphoro-(2-methylimidazole) (2-MeImpG) and uridine-5-phosphoro-(2-methylimidazole) (2-MeImpU), facilitates the incorporation of uridine (U) into oligomeric products with low efficiency. If 2-aminoadenosine (aA) is substituted for adenosine in the template, U is incorporated into the products with much higher efficiency. Random copolymers of C and U act as templates for the efficient synthesis of oligomers from 2-MeImpG and 2-MeImpA only if the concentration of substrates is relatively high (0.1 M). The substitution of 2-MeImpaA permits the reaction to occur with much lower substrate concentrations. This effect is most prominent for template containing large amounts of U.  相似文献   

4.
Na-montmorillonite prepared from Volclay by the titration method facilitates the self-condensation of ImpA, the 5'-phosphorimidazolide derivative of adenosine. As was shown by AE-HPLC analysis and selective enzymatic hydrolysis of products, oligo(A)s formed in this reaction are 10 monomer units long and contain 67% 3',5'-phosphodiester bonds (Ferris and Ertem, 1992a). Under the same reaction conditions, 5'-phosphorimidazolide derivatives of cytidine, uridine and guanosine also undergo self-condensation producing oligomers containing up to 12-14 monomer units for oligo(C)s to 6 monomer units for oligo(G)s. In oligo(C)s and oligo(U)s, 75-80% of the monomers are linked by 2',5'-phosphodiester bonds. Hexamer and higher oligomers isolated from synthetic oligo(C)s formed by montmorillonite catalysis, which contain both 3',5'- and 2',5'-linkages, serve as catalysts for the non-enzymatic template directed synthesis of oligo(G)s from activated monomer 2-MeImpG, guanosine 5'-phospho-2-methylimidazolide (Ertem and Ferris, 1996). Pentamer and higher oligomers containing exclusively 2',5'-linkages, which were isolated from the synthetic oligo(C)s, also serve as templates and produce oligo(G)s with both 2',5'- and 3',5'-phosphodiester bonds. Kinetic studies on montmorillonite catalyzed elongation rates of oligomers using the computer program SIMFIT demonstrated that the rate constants for the formation of oligo(A)s increased in the order of 2-mer < 3-mer < 4-mer ... < 7-mer (Kawamura and Ferris, 1994). A decameric primer, dA(pdA)8pA bound to montmorillonite was elongated to contain up to 50 monomer units by daily addition of activated monomer ImpA to the reaction mixture (Ferris, Hill and Orgel, 1996). Analysis of dimer fractions formed in the montmorillonite catalyzed reaction of binary and quaternary mixtures of ImpA, ImpC, 2-MeImpG and ImpU suggested that only a limited number of oligomers could have formed on the primitive Earth rather than equal amounts of all possible isomers (Ertem and Ferris, 2000). Formation of phosphodiester bonds between mononucleotides by montmorillonite catalysis is a fascinating discovery, and a significant step forward in efforts to find out how the first RNA-like oligomers might have formed in the course of chemical evolution. However, as has been pointed out in several publications, these systems should be regarded as models rather than a literal representation of prebiotic chemistry (Orgel, 1998; Joyce and Orgel, 1999; Schwartz, 1999).  相似文献   

5.
Poly(2'-deoxyadenosine) and poly(thymidine) constructed of carbonate linkages were synthesized by polycondensation between silyl ether and carbonylimidazolide at the 3'- and 5'-positions of the 2'-deoxyribonucleoside monomers. The N-benzoyl-2'-deoxyadenosine monomer afforded the corresponding polycarbonate together with the cyclic oligomers. However, the deprotection of the N-benzoyl group resulted in the scission of the polymer main chain. Thus, the N-unprotected 2'-deoxyadenosine monomers were examined for polycondensation. However, there was involved the undesired reaction between the adenine amino group and the carbonylimidazolide to form the carbamate linkage. In order to exclude this unfavorable reaction, dynamic protection was employed. Strong hydrogen bonding was used in place of the usual covalent bonding for reducing the nucleophilicity of the adenine amino group. Herein, 3',5'-O-diacylthymidines that form the complementary hydrogen bonding with the adenine amino group were added to the polymerization system of the N-unprotected 2'-deoxyadenosine monomer. Consequently, although the oligomers (M(n) = 1000-1500) were produced, the contents of the carbamate group were greatly reduced. The dynamic protection reagents were easily and quantitatively recovered as the MeOH soluble parts from the polymerization mixtures. In the polycondensation of the thymidine monomer, there tended to be involved another unfavorable reaction of carbonate exchange, which consequently formed the irregular carbonate linkages at not only the 3'-5' but also the 3'-3' and 5'-5' positions. Employing the well-designed monomer suppressed the carbonate exchange reaction to produce poly(thymidine) with the almost regular 3'-5'carbonate linkages.  相似文献   

6.
Summary Selected imidazolide-activated nucleotides have been subjected to hydrolysis under conditions similar to those that favor their template-directed oligomerization. Rate constants of hydrolysis of the P–N bond in guanosine 5-monophosphate 2-methylimidazolide (2-MeImpG) and in guanosine 5-monophosphate imidazolide (ImpG), kh, have been determined in the presence/absence of magnesium ion as a function of temperature and polycytidylate [poly(C)] concentration. Using the rate constant of hydrolysis of 2-MeImpG and the rate constant of elongation, i.e., the reaction of an oligoguanylate with 2-MeImpG in the presence of poly(C) acting as template, the limiting concentration of 2-MeImpG necessary for oligonucleotide elongation to compete with hydrolysis can be calculated. The limiting concentration is defined as the initial concentration of monomer that results in its equal consumption by hydrolysis and by elongation. These limiting concentrations of 2-MeImpG are found to be 1.7 mM at 37°C and 0.36 mM at 1°C. Boundary conditions in the form of limiting concentration of activated nucleotide may be used to evaluate a prebiotic model for chemical synthesis of biopolymers. For instance, the limiting concentration of monomer can be used as a basis of comparison among catalytic, but nonenzymatic, RNA-type systems.We also determined the rate constant of dimerization of 2-MeImpG, k2=0.45±0.06 M–1 h–1 in the absence of poly(C), and 0.45±0.06k20.97±0.13 M–1 h–1 in its presence at 37°C and pH 7.95. This dimerization, as well as the trimerization of 2-MeImpG, which represent the first steps in the oligomerization reaction, are markedly slower than the elongation of longer oligoguanylates, (pG) n n>6. This means that in the presence of low concentrations of 2-MeImpG (1.7 mM) the system directs the elongation of longer oligomers more efficiently than the formation of short oligomers such as dimers and trimers. These results will be discussed as a possible example of chemical selection in template-directed reactions of nucleotides.  相似文献   

7.
Phosphoimidazolide activated ribomononucleotides (*pN) are useful substrates for the non-enzymatic synthesis of polynucleotides. However, dilute neutral aqueous solutions of *pN typically yield small amounts of dimers and traces of polymers; most of *pN hydrolyzes to yield nucleoside 5-monophosphate. Here we report the self-condensation of nucleoside 5-phosphate 2-methylimidazolide (2-MeImpN with N = cytidine, uridine or guanosine) in the presence of Mg2+ in concentrated solutions, such as might have been found in an evaporating lagoon on prebiotic Earth. The product distribution indicates that oligomerization is favored at the expense of hydrolysis. At 1.0 M, 2-MeImpU and 2-MeImpC produce about 65% of oligomers including 4% of the 3,5-linked dimer. Examination of the product distribution of the three isomeric dimers in a self-condensation allows identification of reaction pathways that lead to dimer formation. Condensations in a concentrated mixture of all three nucleotides (U,C,G mixtures) is made possible by the enhanced solubility of 2-MeImpG in such mixtures. Although percent yield of internucleotide linked dimers is enhanced as a function of initial monomer concentration, pyrophosphate dimer yields remain practically unchanged at about 20% for 2-MeImpU, 16% for 2-MeImpC and 25% of the total pyrophosphate in the U,C,G mixtures. The efficiency by which oligomers are produced in these concentrated solutions makes the evaporating lagoon scenario a potentially interesting medium for the prebiotic synthesis of dimers and short RNAs.  相似文献   

8.
Uracil DNA glycosylase excises uracil residues from DNA that can arise as a result of deamination of cytosine or incorporation of dUMP residues by DNA polymerase. We have carried out a detailed study to define the specificities and the kinetic parameters for its substrates by using a number of synthetic oligodeoxyribonucleotides of varying lengths and containing uracil residue(s) in various locations. The results show that the Escherichia coli enzyme can remove a 5'-terminal U from an oligomer only if the 5'-end is phosphorylated. The enzyme does not remove U residues from a 3'-terminal position, but U residues can be excised from oligonucleotides with either pd(UN)p or pd(UNN) 3'-termini. The oligomer d(UUUUT) can have the second or third U residues from the 5'-end excised even when the neighboring site is an abasic site (3' or 5', respectively). On the basis of these findings, pd(UN)p was anticipated to be the smallest size substrate. Results show detectable amounts of U release from the substrate pd(UT)p; however, significantly higher amounts of U release were observed from pd(UT-sugar) or pd(UTT). Determinations of the Km and Vmax values show that the different rates of U excision from oligomers of different sizes (trimeric to pentameric) but containing U in the same position are largely due to the differences in the Km values, whereas the different rates of U excision from the substrates of the same size but containing U in different positions are largely due to different Vmax values.  相似文献   

9.
The processing of a unique uracil in DNA has been studied in mammalian cells. A synthetic oligodeoxyribonucleotide carrying a potential Bgl II restriction site, where one base has been substituted with a uracil, was inserted in the early intron of SV40 genome. Various heteroduplexes were constructed in such a manner that the restitution of an active Bgl II restriction site corresponds in each case to the specific substitution of the uracil by one of the four bases normally present in the DNA. DNA cuts by this restriction enzyme in one or several constructed heteroduplexes immediately determine the type of base pair substitution produced at the site of the U residue. When the uracil is inserted opposite a purine it is fully repaired; when facing a guanine it is replaced by a cytosine and opposite an adenine it is replaced by a thymine. These results indicate the error-free repair of uracil when it appears in the cell with the usual mechanisms such as cytosine deamination or incorporation of dUTP in place of dTTP during replication. When the uracil is inserted opposite a pyrimidine no error free repair at all is detected for U:C or U:T mismatches. It appears, moreover, that in approximately 18% of the cases U:T mismatch leads to a C:G base pairing. In the majority of the U:pyrimidine mismatches, mutations occur in the vicinity of the uracil, including base substitutions and frameshifts by addition of one or several bases.  相似文献   

10.
Zeng Y  Wang Y 《Biochemistry》2007,46(27):8189-8195
Nucleoside 5-bromo-2'-deoxyuridine (BrdU), after being incorporated into cellular DNA, is well-known to sensitize cells to ionizing radiation and UV irradiation. We reported here, for the first time, the sequence-dependent formation of intrastrand cross-link products from the UVB irradiation of BrdU-treated MCF-7 human breast cancer cells. Our results showed that BrdU replaced more than 30% dT in genomic DNA after the cells were treated with 10 microM BrdU for 48 h. LC-MS/MS data revealed that more than 50% of the incorporated BrdU was consumed during UVB irradiation, of which more than half was dehalogenated to yield dU. Low-dose (5.0 kJ/m2) UVB irradiation of BrdU-treated cells yielded four intrastrand cross-link products, where the C5 of uracil is covalently bonded to the C8 of its neighboring 5' or 3' guanine to give G[8-5]U and U[5-8]G, respectively, and the C5 of uracil could couple with the C2 or C8 of its vicinal 5' adenine to give A[2-5]U and A[8-5]U, respectively. All the above cross-link products except A[2-5]U could also be induced in BrdU-treated cells upon UVB irradiation at a dose of 39 kJ/m2. We further demonstrated, by using LC-MS/MS, that the yield of G[8-5]U was much greater than the total yields of A[2-5]U and A[8-5]U. In addition, our results revealed that BrdU treatment stimulated considerably the UVB-induced formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) in vivo. The formation of these intrastrand cross-link products and 8-oxo-dG in vivo underscores the importance of these products in the photosensitizing effect of BrdU.  相似文献   

11.
(Guanosine 5′-phosphor)-2-methylimidazolide (2-MeImpG), unlike guanosine 5′-phosphorimidazolide (ImpG), undergoes an efficient, buffer-independent, template-directed oligomerization in the presence of poly(C) at pH values above 7.6. The reaction occurs in a Watson-Crick double helix and yields predominantly 3′-5′-linked oligomers up to the 50-mer in above 80% yield. Synthesis proceeds in the 5′ → 3′ direction and has high fidelity in the sense that nucleotides other than G are not incorporated significantly into oligomers. Under some conditions, oligomers corresponding to approximately one and two turns of the helix are obtained in higher yield than somewhat longer or somewhat shorter oligomers.In the protonated triple-helical structure formed below pH 7, the efficiency of the oligomerization is much lower. Oligomers up to about the 10-mer are obtained. The most abundant products are “capped” at the 5′ terminus with a GppG pyrophosphate group.  相似文献   

12.
Summary A computer simulation (KINSIM) modeling up to 33 competing reactions was used in order to investigate the product distribution in a template-directed oligonucleotide synthesis as a function of time and concentration of the reactants. The study is focused on the poly(C)-directed elongation reaction of an oligoguanylate (a 7-mer is chosen) with guanosine 5-monophosphate-2-methylimidazolide (2-MeImpG), the activated monomer. It is known that theelongation of oligoguanylates to form oligomeric products such as 8-mer, 9-mer, 10-mer, etc., is in competition with (1) thedimerization and further oligomerization reaction of 2-MeImpG that leads to the formation of dimers and short oligomers, and (2) thehydrolysis of 2-MeImpG that forms inactive guanosine 5-monophosphate, 5-GMP. Experimentally determined rate constants for the above three processes at 37°C and pH 7.95 were used in the simulation; the initial concentrations of 2-MeImpG, [M]o, and of the oligoguanylate primer, [7-mer]o, were varied, and KINSIM calculated the distribution of products as a function of time until equilibration was reached, i.e., when all the activated monomer has been consumed. In order to sort out how strongly the elongation reaction may be affected by the competing hydrolysis and dimerization, we also simulated the idealized situation in which these competing reactions do not occur. Simulation of the idealized system suggests that (1) the fraction of [7-mer]o that has reacted as well as the product distribution after equilibration do not depend on the absolute concentrations of the reactants, but only on their ratio, [M]o/[7-mer]o; (2) the rate of elongation is proportional to [7-mer]o and not to [M]o; and (3) as the [M]o/[7-mer]o ratio increases longer oligomers are formed. The results of the computer simulation with the experimental system, i.e., elongation in the presence of both hydrolysis and dimerization, are similar to the ones obtained with the idealized system as long as dimerization and hydrolysis are not responsible for consuming a substantial fraction of 2-MeImpG.  相似文献   

13.
Sung JS  Mosbaugh DW 《Biochemistry》2000,39(33):10224-10235
Escherichia coli double-strand uracil-DNA glycosylase (Dug) was purified to apparent homogeneity as both a native and recombinant protein. The molecular weight of recombinant Dug was 18 670, as determined by matrix-assisted laser desorption-ionization mass spectrometry. Dug was active on duplex oligonucleotides (34-mers) that contained site-specific U.G, U.A, ethenoC.G, and ethenoC.A targets; however, activity was not detected on DNA containing a T.G mispair or single-stranded DNA containing either a site-specific uracil or ethenoC residue. One of the distinctive characteristics of Dug was that the purified enzyme excised a near stoichiometric amount of uracil from U.G-containing oligonucleotide substrate. Electrophoretic mobility shift assays revealed that the lack of turnover was the result of strong binding by Dug to the reaction product apyrimidinic-site (AP) DNA. Addition of E. coli endonuclease IV stimulated Dug activity by enhancing the rate and extent of uracil excision by promoting dissociation of Dug from the AP. G-containing 34-mer. Catalytically active endonuclease IV was apparently required to mediate Dug turnover, since the addition of 5 mM EDTA mitigated the effect. Further support for this interpretation came from the observations that Dug preferentially bound 34-mer containing an AP.G target, while binding was not observed on a substrate incised 5' to the AP-site. We also investigated whether Dug could initiate a uracil-mediated base excision repair pathway in E. coli NR8052 cell extracts using M13mp2op14 DNA (form I) containing a site-specific U.G mispair. Analysis of reaction products revealed a time dependent appearance of repaired form I DNA; addition of purified Dug to the cell extract stimulated the rate of repair.  相似文献   

14.
Uracyl and adenine containing oligomers derived from carboxymethyl derivatives of morpholine nucleoside analogues (MorGly) were synthesized using the methods of peptide chemistry. Capillary electrophoresis conditions were found for the analysis of the homogeneity of the nucleic acid mimics protonated at physiological pH. The thermal stability of complementary complexes formed by the MorGly oligomers was shown to depend dramatically on the heterocyclic base structure (uracil or adenine). Based on the study of tandem complexes it was demonstrated that the impact on the thermal stability of cooperative interactions at oligomer junctions was higher for modified oligomers than for native oligodeoxyriboadenylates. Adenine containing MorGly oligomers formed more stable complexes with poly(U) than native oligodeoxyriboadenylates of the same length. Complexes formed by modified oligomers with polyribonucleotides were more stable if compared with polydeoxyribonucleotides.  相似文献   

15.
Aqueous N2O/O2-saturated solutions of poly(U) were irradiated at 0 degrees C and the release of unaltered uracil determined. Immediately after irradiation G(uracil release) was 1.5 which increased to a value of 5.3 +/- 0.3 upon heating to 95 degrees C. Thereby all of the organic hydroperoxides (G = 6.8 +/- 0.7) and some of the hydrogen peroxide (G = 1.7 +/- 0.2) was destroyed leaving G(peroxidic material; mainly hydrogen peroxide) = 1.0 +/- 0.7. G(chromophore loss) = 8-11 was measured immediately after irradiation, but no increase was observed upon heating. Addition of iodide destroyed the hydroperoxides and caused immediate base release to rise to G = 4 and further heating brought the value to that observed in the absence of iodide. In contrast, on reducing the hydroperoxides with NaBH4, immediate uracil release rose to only G = 2.8 and no further increase was observed on heating. A major product (G = 2.7) is carbon dioxide. There are also osazone-forming compounds produced (G = 2.7), all of which are originally bound to poly(U). Heating in acid solutions, as is required for this test, releases glycoladehyde-derived osazone (G = 0.8) and further unidentified low molecular weight material (G = 0.9). It is concluded that the primary radicals which cause these lesions are the base OH adduct radicals. In the presence of oxygen these are converted into the corresponding peroxyl radicals which abstract an H atom from the sugar moiety. In the course of this reaction base-hydroperoxides are formed. However, such base hydroperoxides cannot be the only organic hydroperoxides, but some (G congruent to 2.5) sugar-hydroperoxides must be formed as indicated by the increase in base release by the addition of iodide. It is speculated that a sugar-hydroperoxide located at C(3') is reduced by iodide to a carbonyl function at C(3'), a lesion that releases the base, while reduction with NaBH4 reduces it to an alcohol function at C(3') thus preventing base release.  相似文献   

16.
DNA with abasic sites has been prepared by deamination of cytosine followed by treatment of the product with uracil N-glycosylase. Termination in vitro on such templates does not occur until treatment with uracil N-glycosylase. DNA terminated one base before abasic sites created from C's has been used as a template in "second stage" reactions. With enzymes devoid or deficient in 3' greater than 5' exonuclease activity purines, particularly adenine, are preferentially added opposite the putative abasic site. 2-Aminopurine behaves more like adenine than like guanine in these experiments. Polymerase beta preferentially incorporates A opposite abasic sites produced from T, and G opposite abasic sites produced from C. We have eliminated an obvious artefact (e.g. strand switching) which might account for this observation.  相似文献   

17.
The crystal structure of the RNA octamer, 5'-GGCGUGCC-3' has been determined from x-ray diffraction data to 1.5 angstroms resolution. In the crystal, this oligonucleotide forms five self-complementary double-helices in the asymmetric unit. Tandem 5'GU/3'UG basepairs comprise an internal loop in the middle of each duplex. The NMR structure of this octameric RNA sequence is also known, allowing comparison of the variation among the five crystallographic duplexes and the solution structure. The G.U pairs in the five duplexes of the crystal form two direct hydrogen bonds and are stabilized by water molecules that bridge between the base of guanine (N2) and the sugar (O2') of uracil. This contrasts with the NMR structure in which only one direct hydrogen bond is observed for the G.U pairs. The reduced stability of the r(CGUG)2 motif relative to the r(GGUC)2 motif may be explained by the lack of stacking of the uracil bases between the Watson-Crick and G.U pairs as observed in the crystal structure.  相似文献   

18.
Template-directed synthesis on the oligonucleotide d(C7-G-C7)   总被引:1,自引:0,他引:1  
When the deoxynucleotide template d(C7-G-C7) is incubated with the activated nucleotides 2-MeImpG and 2-MeImpC, a series of oligomers of G up to the sevenmer and a series of copolymers of composition GnC with n = 3 to 13 are formed. Oligomers GnC with n greater than 7 are completely degraded by pancreatic ribonuclease, establishing that they contain a 3' to 5' internucleotide bond between 5'-C and 3'-G within a sequence of the form (pG)ipC(pG)j. As expected, (pG)7-Cp and (pG)6-Cp are major hydrolysis products. Detailed analysis of the product distribution shows that a substantial fraction of the oligomeric products are of the type (pG)ipC(pG)j with i less than 7. This shows that product synthesis does not necessarily begin at the 3' terminus of the template. The significance of this finding in terms of the origin of molecular replication is discussed.  相似文献   

19.
The synthesis of a diaminopurine PNA monomer, N-[N6-(benzyloxycarbonyl)-2,6-diaminopurine-9-yl] acetyl-N-(2-t-butyloxycarbonylaminoethyl)glycine, and the incorporation of this monomer into PNA oligomers are described. Substitution of adenine by diaminopurine in PNA oligomers increased the T m of duplexes formed with complementary DNA, RNA or PNA by 2.5-6.5 degrees C per diaminopurine. Furthermore, discrimination against mismatches facing the diaminopurine in the hybridizing oligomer is improved. Finally, a homopurine decamer PNA containing six diaminopurines is shown to form a (gel shift) stable strand displacement complex with a target in a 246 bp double-stranded DNA fragment.  相似文献   

20.
G · C complex formation was studied by infrared spectroscopy for a system that has been shown by Inoue & Orgel (1982) to give efficient, template-dependent synthesis of oligo(G). Guanosine-5′-phosphor-2-methylimiazolide (2-MeImpG) exhibits rapid formation with poly(C) of a G · C double helix at pD ~ 8 and of a C · G · CH+ triple helix at pD ~ 6.5 in the presence of Na+. Significant oligo(G) synthesis does not occur under these conditions. In the presence of synthetically effective concentrations of Mg2+ G · C complex formation is much slower but eventually goes to completion. The rate of complex formation parallels that of chemical synthesis. Infrared spectra and melting curves confirm that oligo(G) of high molecular weight is formed in high yield. The bulk of the G · C complex at any given time during the reaction is composed of G residues that have already been polymerized and not of the monomer 2-MeImpG. Evidence indicates that synthesis proceeds primarily at growing points at the ends of the G · C helical regions and not randomly on a fully occupied template.  相似文献   

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