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PubMed1985

[Determination of the adenine-specific methylase recognition site of Shigella sonnei 47, using hydrazinolysis with subsequent separation of purine oligonucleotides by thin layer chromatography on DEAE- cellulose].

A procedure for separation of oligopurine blocks of different length and composition by two-dimensional thin layer chromatography on DEAE-cellulose plates has been developed. This method allows a comparative analysis of the purine isostich content in the DNAs of various origin. In case of methylated DNA, the method permits to compare the substrate specificity of different enzymes responsible for the adenine residue methylation in the DNA. In combination with enzymatic treatment of labeled methylated isostichs, the method described can be used for the deciphering of the methylated sequences as well as for constructing, in a number of cases, the recognition site of adenine-specific methylases. Thus, it was demonstrated that methylase SsoI recognizes the 5...G-A-A-T-T-C ... 3' sequence and methylates its adenine residue nearest to the 5'-end.

Adenine↗
PubMed2000

Secondary binding sites for triplex-forming oligonucleotides containing bulges, loops, and mismatches in the third strand.

We have used DNase I footprinting to examine the binding of five different 17-mer oligonucleotides to a 53-base oligopurine tract containing four pyrimidine interruptions. Although all the expected triplexes formed with high affinity (K(d) approximately 10-50 nM), one oligonucleotide produced a footprint at a second site with about 20-fold lower affinity. We have explored the nature of this secondary binding site and suggest that it arises when each end of the third strand forms a 7-mer triplex with adjacent regions on the duplex, generating a contiguous 14-base triplex with a bulge in the center of the third strand oligonucleotide. This unusual binding mode was examined by use of oligonucleotides that were designed with the potential to form different length third-strand loops of various base composition. We find that triplexes containing single-base bulges are generally more stable than those with dinucleotide loops, though triplexes can be formed with loops of up to nine thymines, generating complexes with submicromolar dissociation constants. These structures are much more stable than those formed by adding two separate 7-mer oligonucleotides, which do not generate DNase I footprints, though a stable complex is generated when the two halves are covalently joined by a hexa(ethylene glycol) linker. MPE produces less clear footprints, presumably because this cleavage agent binds to triplex DNA, but confirms that the oligonucleotides can bind in unexpected places. These results suggest that extra care needs to be taken when designing long triplex-forming oligonucleotides so as to avoid triplex formation at shorter secondary sites.

Base Sequence↗
PubMed2001

Characterization of oligonucleotide/lipid interactions in submicron cationic emulsions: influence of the cationic lipid structure and the presence of PEG-lipids.

We have recently described how oligonucleotide (ON) stability and release from O/W cationic emulsions are governed by the lipid composition. The aim of the present paper was to investigate the properties of the ON/lipid complexes through fluorescence resonance energy transfer (FRET), size, surface tension measurements and cryomicroscopy. Starting from a typical emulsion containing stearylamine as a cationic lipid, the influence of the lipid structure (monocationic molecules bearing mono or diacyl chains, or polycations) as well as of the presence of PEGylated lipids, were studied. The presence of a positive charge on the droplet surface clearly contributed to enhance the ON interaction with lipid monolayers and to bring the ON molecules closer to the interface. Hydrophobic interactions through the acyl chains were shown to further enhance the anchorage of the ON/lipid complexes. In contrast, the incorporation of PEGylated lipids acted as a barrier against the establishment of electrostatic bindings, the polyethyleneglycol chains acting themselves as interaction sites for the ON leading to hydrophilic complexes. Similar features were observed for the polycationic lipid, and cryomicroscopy revealed the existence of bridges of various intensities between the droplets of the emulsion containing either PEG or the polycation, probably because of the configuration of the ON at the interface.

Algorithms↗
PubMed2005

Discrimination between perfect and mismatched duplexes with oligonucleotide gel microchips: role of thermodynamic and kinetic effects during hybridization.

The efficiency of discrimination between perfect and mismatched duplexes during hybridization on microchips depends on the concentrations of target DNA in solution and immobilized probes, buffer composition, and temperature of hybridization and is determined by both thermodynamic relationships and hybridization kinetics. In this work, optimal conditions of discrimination were studied using hybridization of fluorescently labeled target DNA with custom-made gel-based oligonucleotide microchips. The higher the concentration of immobilized probes and the higher the association constant, the higher the concentration of the formed duplexes and the stronger the corresponding fluorescence signal, but, simultaneously, the longer the time needed to reach equilibrium. Since mismatched duplexes hybridize faster than their perfect counterparts, perfect-to-mismatch signal ratio is lower in transient regime, and short hybridization times may hamper the detection of mutations. The saturation time can be shortened by decreasing the probe concentration or augmenting the gel porosity. This improves the detection of mutations in transient regime. It is shown that the decrease in the initial concentration of oligonucleotide probes by an order of magnitude causes only 1.5-2.5-fold decrease of fluorescence signals after hybridization of perfect duplexes for 3-12 h. At the same time, these conditions improve the discrimination between perfect and mismatched duplexes more than two-fold. A similar improvement may be obtained using an optimized dissociation procedure.

Base Composition↗
PubMed2001

Fluorescence fluctuation analysis for the study of interactions between oligonucleotides and polycationic polymers.

The interactions between a cationic polymer, poly(2-dimethylamino)ethyl methacrylate (pDMAEMA), and negatively charged rhodamine-labeled 25-mer phosphodiester oligonucleotides (Rh-ONs) were studied by fluorescence fluctuation spectroscopy and other techniques. The composition of the pDMAEMA/Rh-ON complexes was investigated as a function of the charge ratio (+/-) by increasing the pDMAEMA concentration and keeping the Rh-ON concentration constant. We applied two different methods for analyzing the fluorescence fluctuation profiles of the pDMAEMA/Rh-ON complexes, which depended on their composition. First, we analyzed the data with the photon counting histogram (PCH) technique, which determines the molecular brightness and the concentration of fluorophores (Chen et al, 1999). A particular challenge for the data analysis is the occurrence of sudden fluorescence bursts in the fluorescence fluctuation profiles, which are linked to the appearance of multimolecular complexes (i. e. when several Rh-ONs were present in one complex). A quantitative interpretation of the analysis for the complexes remains challenging and is connected to the rarity of the fluorescent bursts, which do not provide sufficient data statistics. To specifically address the problem of the fluorescent bursts we employed a method described by Van Craenenbroeck et al. (1999). This method, applicable only when data were integrated over much longer time bins, allowed us to estimate the number of fluorescence bursts which could be considered as a relative measure of the amount of multimolecular complexes present. When monomolecular complexes were formed, i. e. at high values of the charge ratio, highly intense fluorescence peaks were not present and the interpretation of the PCH analysis was more straightforward. The molecular brightness of the species (epsilon), as revealed from PCH analysis, was greater than epsilon for the free Rh-ONs, indicating that the Rh-ONs were attached to pDMAEMA chains.

Cations↗
PubMed1996

The structure of B-DNA in oriented fibers.

Native, general sequence B-form DNA in uniaxially oriented fibers is a ten-fold helix with identical antiparallel strands: this is to say the molecular symmetry is 2 2 10(1). The diffraction patterns indicate that local variations, however significant, must be modest. This is true also for the lithium salt of calf thymus DNA in fibers that are polycrystalline as well as oriented. The contents of its orthorhombic unit cells are arranged with P2(1)2(1)2(1) symmetry which permits the molecular symmetry to be merely two-fold. The molecular structure of DNA in such conditions resembles, conformationally and molecularly, that of B-type DNA in oligonucleotide single crystals and in oriented polycrystalline fibers of polyoligonucleotides, and therefore provides a basis for evaluating the variations that may be due to sequence effects in polyoligonucleotides in fibers and oligonucleotides in single crystals.

Base Composition↗
PubMed2006

CpG 7909: PF 3512676, PF-3512676.

CpG 7909 [PF-3512676] is an immunomodulating synthetic oligonucleotide designed to specifically agonise the Toll-like receptor 9 (TLR9). It is being developed for the treatment of cancer [ProMune] as a monotherapy and in combination with chemotherapeutic agents, and it is also under development as an adjuvant [VaxImmune] for vaccines against cancer and infectious diseases. CpG 7909, acting through the TLR9 receptor present in B cells and plasmacytoid dendritic cells, stimulates human B-cell proliferation, enhances antigen-specific antibody production and induces interferon-alpha production, interleukin-10 secretion and natural killer cell activity. Coley Pharmaceutical Group originally developed CpG 7909 using its CpG DNA technology. In March 2005, Coley granted Pfizer an exclusive global license to develop and commercialise CPG 7909 [ProMune] for the treatment, control and prevention of multiple cancer indications. Coley licensed CpG 7909 [VaxImmune] to Chiron Corporation for adjuvant use with Chiron's prophylactic vaccine candidates against infectious diseases in December 2003. Chiron was acquired by and merged into Novartis in April 2006. In 2002, GlaxoSmithKline (GSK) was granted a worldwide, non-exclusive licence to Coley's CpG immunostimulatory oligonucleotides, including CpG 7909 [VaxImmune], for their use as adjuvants for cancer vaccines. In 2000, Coley entered into a co-exclusive licensing agreement with GSK for the development of therapeutic and prophylactic vaccines against infectious diseases. This licensing agreement included CpG 7909 [VaxImmune] and other CpG-based immunostimulatory oligonucleotides. In September 2004, Coley Pharmaceuticals was awarded a 16.9 million US dollars, 5-year contract from the National Institute of Allergy and Infectious Diseases (NIAID), one of the National Institutes of Health (NIH), to support the development of novel immune-activating drugs for defense against bioterror agents. This contract will be used to expand Coley's proprietary line of TLR Therapeutic products. Together with prior awards, the new contract brings the total committed biodefense funding for Coley to 35 million US dollars. During the first quater of 2006, Pfizer disclosed its intention to develop CpG 7909 for breast cancer. A phase I/II trial in patients with NHL has also been conducted in 24 patients with relapsed or refractory disease at the University of Iowa. Pfizer initiated two international phase III trials under the special protocol assessment (SPA) procedure of the US FDA. These trials are evaluating CpG 7909 in combination with chemotherapy versus chemotherapy alone as a first-line treatment for patients with advanced (stage IIIb or IV) non-small-cell lung cancer (NSCLC). Approximately 800 patients will be enrolled in each trial. The primary endpoint is overall survival time. In 2005, Coley completed a multinational phase IIb trial of CpG 7909 in combination with chemotherapy in 112 patients with NSCLC. The goal of the study was to improve the outcome of standard first-line chemotherapeutic regimens (taxane and platinum) for NSCLC by adding CpG 7909. Coley has been granted 11 US patents, covering key aspects of the company's CpG TLR9 antagonist technologies. Three US patents relating to CpG 7909 and the use of certain oligonucleotides for treating cancer are due to expire in 2014. Coley also has pending US patent applications covering the specific sequence of CpG 7909 and its use to treat cancer, which, if issued, would be expected to expire between 2014 and 2017. In April 2004, Coley received US Patent No. 6,727,230 covering the use of oligonucleotides containing at least one phosphorothioate linkage to stimulate cellular immune responses. In November 2003, Coley received US Patent No. 6,653,292, which protects the use of TLR9-containing oligonucleotides to treat or prevent cancer or to enhance multimodal cancer treatment regimens. In July 2002, Coley received US Patent No. 6,406,705 covering its CpG oligonucleotide immunostimulants for use in combination with conventional adjuvants. In April 2001, Coley was issued with US Patent No. 6,207,646 covering the composition and use of its immune stimulants including CpG 7909. In September 2001, Coley was granted US Patent No. 6,214,806, which expands the coverage on the use of CpG oligonucleotides in the treatment of certain respiratory disorders.

Adjuvants, Immunologic↗
PubMed1991

Synthesis of oligonucleotide derivatives with dansyl group at sugar fragment.

The method for introduction of dansyl group into the 2'-position of oligonucleotides is described. The dansyl group was introduced to 2'-position of uridine by the reaction of dansyl chloride with 2'-amino-2'-deoxyuridine, which was then converted to the 5'-dimethoxytrityl phosphorobisdiethylamidite derivative, (4). This reagent was used for the solid-phase synthesis of the oligonucleotide-dansyl conjugate, 5'-ACTCU(DNS)AGAGG. The conjugate could be purified by reversed-phase HPLC. The nucleoside composition of the conjugate was verified by the enzymatic digestion analysis.

Base Sequence↗
PubMedDate not supplied

[Analysis of perfect and mismatched DNA duplexes by a generic hexanucleotide microchip].

The thermodynamic analysis was done for the duplexes formed by fluorescently labeled oligonucleotide targets on a genetic hexanucleotide microchip. All 4096 different hexanucleotide chains were immobilized as probes in individual gel pads of the microchip. To strengthen the hybridization, each hexamer was extended at both ends by one nucleotide from the equimolar mixture of all four nucleotides to serve as nonselective linkers. It has been shown that the melting curves for oligonucleotide duplexes formed on the microchip and in a solution are quite similar. The influence of ionic surrounding has been studied in terms of the hybridization efficiency and discrimination between the mismatched and perfect duplexes. Different approaches have been tested to compensate the dependence of duplex stability on the GC content. It has been demonstrated that the use of chaotropic agents, addition of nonlabeled GC-rich competitor oligonucleotides, as well as creation of a temperature gradient along the microchip reproducing the distribution of melting temperatures, efficiently level out the AT/GC differences. The use of tetramethylammonium chloride for the same purpose was accompanied by weakening to some extent the discrimination between the mismatched duplexes and the perfect ones.

Base Composition↗
PubMed2006

Topoisomerase action on short DNA duplexes reveals requirements for gate and transfer DNA segments.

Type II topoisomerases change DNA topology by passage of one DNA duplex (the transfer, T-segment) through a transient double-stranded break in another (the gate, G-segment). Here we monitor the passage between short double-stranded DNA segments within long single-stranded DNA circles that leads to catenation of the circles. To facilitate catenation, the circles were brought into close proximity using a tethering oligonucleotide, which was removed after the reaction was complete. We varied the length and the composition of the reacting DNA segments. The minimal DNA duplex length at which we detected catenation was 50-60 bp for DNA gyrase and 40 bp for topoisomerase IV (Topo IV). For Topo IV, catenation was observed when one, but not both, of the DNA-DNA duplexes was replaced by a DNA-RNA duplex. Topo IV cleaved the DNA-DNA duplex, but not the DNA-RNA duplex implying that the DNA-RNA duplex can be a T-segment but not a G-segment.

Adenosine Triphosphate↗
PubMed1987

Molecular cloning of cDNA coding for rat proliferating cell nuclear antigen (PCNA)/cyclin.

The 'proliferating cell nuclear antigen' (PCNA), also known as cyclin, appears at the G1/S boundary in the cell cycle. Because of its possible relationship with cell proliferation, PCNA/cyclin has been receiving attention. PCNA/cyclin is a non-histone acidic nuclear protein with an apparent mol. wt of 33000-36000. The amino acid composition and the sequence of the first 25 amino acids of rabbit PCNA/cyclin are known. Using an oligonucleotide probe corresponding to the sequence of the first five amino acids, a cDNA clone for PCNA/cyclin was isolated from rat thymocyte cDNA library. The cDNA (1195 bases) contains an open reading frame of 813 nucleotides coding for 261 amino acids. The 3'-non-coding region is 312 nucleotides long and contains three putative polyadenylation signals. The mol. wt of rat PCNA/cyclin was calculated to be 28 748. The deduced amino acid sequence and composition of rat PCNA/cyclin are in excellent agreement with the published data. Using the cDNA probe, two species of mRNA (1.1 and 0.98 kb) were detected in rat thymocyte RNA. Southern blot analysis of total human genomic DNA suggests that there is a single gene coding for PCNA/cyclin. The deduced amino acid sequence of rat PCNA/cyclin has a similarity with that of herpes simplex virus type-1 DNA binding protein.

Amino Acid Sequence↗
PubMed1998

DNA binding of activator protein-1 is increased in human mesangial cells cultured in high glucose concentrations.

Human mesangial cells (HMC) grown in high glucose environments synthesize excessive amounts of extracellular matrix proteins (ECM). The promoter regions of certain ECM genes contain TPA (phorbol ester)-responsive element (TRE) motifs that bind the transcription factor, activator protein-1 (AP-1), a complex of Jun and other phosphoproteins. AP-1 binding to the TRE promoter is regulated by the quantity, composition and post-translational modifications of proteins in the AP-1 complex. We report an increased binding of AP-1 to TRE oligonucleotides in HMC cultured chronically (5 days) in high glucose environments (30 mM d-glucose). This increased binding is not due to differences in the nuclear quantity of AP-1 proteins or in the composition of the AP-1 complex when compared to AP-1 proteins from cells grown in normal glucose (5 mM d-glucose). A 30 mM l-glucose environment also increased AP-1 binding, but to a degree less than d-glucose. The increased AP-1 binding was partly reversed by treatment of HMC with Calphostin C or Bisindolylmaleimide I suggesting a partial role of the protein kinase C (PKC) pathway in mediating AP-1 binding. AP-1 binding was unaffected by treatment of cells with the MEK inhibitor PD 98059. In addition, increased AP-1 binding persisted for at least 48 hours after media glucose concentrations were normalized. The level of Jun-NH2-terminal kinase (JNK) activity and the phosphorylation of the JNK kinase, SEK1, were unchanged by chronic high glucose concentrations. These studies suggest that in HMC cultured in chronic high glucose, post-translational modifications increase the binding of AP-1 to the TRE motif.

Binding Sites↗
PubMed1983

Mass spectrometric monitoring of solid phase phosphate triester synthesis of DNA fragments.

A new and convenient pyrolysis mass spectrometric approach for monitoring solid phase phosphate triester synthesis of oligodeoxyribonucleotides in 5'-3' direction has been developed. The base-typical fragment ions produced by pyrolysis at 280 degrees C of the polymer-bound oligonucleotide triesters in the mass spectrometer permit the analytical monitoring of DNA chain growth, using simple mathematical operations. The base composition can be determined directly from the polymer. In addition, minor nucleosides can easily be detected.

Base Sequence↗
PubMed1999

Peroxynitrite-induced reactions of synthetic oligonucleotides containing 8-oxoguanine.

8-Oxoguanine (8-oxo-G) is one of the most common DNA lesions present in normal tissues due to exposure to reactive oxygen species. Studies at this and other laboratories suggest that 8-oxo-G is highly susceptible to secondary oxidation, making it a likely target for endogenous oxidizing agents, such as peroxynitrite (ONOO-). Synthetic oligonucleotides containing 8-oxoguanine were treated with ONOO-, and the reaction products were analyzed by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI--MS). CCACAACXCAAA, CCAAAGGXAGCAG, CCAAAXGGAGCAG, and TCCCGAGCGGCCAAAGGXAGCAG (X is 8-oxo-G) were found to readily react with peroxynitrite via the same transformations as those observed for free 8-oxo-2'-deoxyguanosine. The composition of the reaction mixtures was a function of ONOO- concentration and of the storage time after exposure. The oligonucleotide products isolated at low [ONOO-]/[DNA] ratios (<5) were tentatively assigned as containing 3a-hydroxy-5-imino-3,3a,4,5-tetrahydro-1H-imidazo[4, 5d]imidazol-2-one, 5-iminoimidazolidine-2,4-dione, and its hydrolytic product, oxaluric acid. At a [ONOO-]/[DNA] ratio of >10, 2,4,6-trioxo[1,3,5]triazinane-1-carboxamidine- and cyanuric acid-containing oligomers were the major products. The exact location of a modified base within a DNA sequence was determined using exonuclease digestion of oligonucleotide products followed by LC/ESI--MS analysis of the fragments. For all 8-oxo-G-containing oligomers, independent of the sequence, the reactions with ONOO- took place at the 8-oxo-G residues. These results suggest that 8-oxo-G, if present in DNA, is rapidly oxidized by peroxynitrite and that oxaluric acid is a likely secondary oxidation product of 8-oxo-G under physiological conditions.

Chromatography, Liquid↗
PubMed1987

Analysis of sequences from the extremely A + T-rich genome of Plasmodium falciparum.

The genome of the human malaria parasite Plasmodium falciparum has an A + T content of about 82%, higher than any other organism whose DNA has been characterized. Computer analysis of 36 kb of available nucleotide sequences from this species showed that the coding regions, with an A + T content of 69.0%, are flanked by more A + T-rich regions of 86.0% A + T. Within the coding sequences, the A/T ratio was 1.68 in the mRNA sense strand, and overall A + T content in the three codon positions increased in the order 1st-2nd-3rd position. Codons with T or especially A in the third position were strongly preferred. Codon usage among individual parasite genes was very similar compared to genes from other species. Dinucleotide frequencies for the parasite DNA were close to those expected for a random sequence with the known base composition, except that the CpG frequency in the coding sequences was low.

Adenine↗
PubMed1983

Initiation, processing and termination of ribosomal RNA from a hybrid 5 S ribosomal RNA gene in a plasmid.

Transformation of an RNA-processing mutant (rne, RNase E-) of Escherichia coli with a recombinant plasmid containing the promoter region of the ribosomal cluster rrnA and portions from the 3' region of the rrnD cluster results in the accumulation of the precursors to 5 S ribosomal RNAs at the permissive as well as that of two full-length transcripts and a processing intermediate at the nonpermissive temperature. The two full-length transcripts start from the two rrnA promoters, which are about 120 nucleotides apart. This plasmid, pJR3 delta, contains an intact 5 S rRNA gene and portions from the 16 S and 23 S rRNA genes. Analysis of the major plasmid-specific RNA species revealed that RNA molecules initiated in vivo from the first promoter (P1) start with pppA, while transcripts from the second promoter (P2) contain either pppG or pppC at their 5' ends. Termination occurs mainly at the first available termination site. Full-length transcripts initiated from both promoters are processed to precursors of 5 S rRNAs in vivo at the permissive temperature, but only about 20% of these transcripts are processed to mature 5 S rRNA. RNA1 and RNA2 (the transcripts initiated from P1 and P2, respectively) and RNA3 (an RNA-processing intermediate containing the entire 5 S region and the 3' end of the transcripts) can be cleaved in vitro by cell extracts of wild type strains resulting in precursor and mature 5 S rRNAs in a reaction that is RNase E dependent but not ribosome dependent. The 5' end of the processed 5 S rRNA can correspond to the 5' end of mature 5 S rRNA or it can contain one to three additional nucleotides.

Base Composition↗
PubMed2004

Quantitative X-ray photoelectron spectroscopy and time-of-flight secondary ion mass spectrometry characterization of the components in DNA.

The great diversity of techniques to synthesize and use DNA microarrays has made them extremely flexible for a variety of applications. This flexibility also has made standardization difficult, leading to problems comparing data from these different systems. In this work, we use the surface science techniques of X-ray photoelectron spectroscopy (XPS) and time-of-flight secondary ion mass spectrometry (TOF-SIMS) to analyze the components of DNA. The atomic ratios of the components of nucleobases, nucleosides, and nucleotides were characterized by XPS. The chemical shifts in the high-resolution XPS spectra allow for their relatively easy resolution. The unique positive and negative ions from the nucleobases, nucleosides, and nucleotides in their TOF-SIMS spectra were identified. This information was used to build a comprehensive table of all of the molecular ions. These standard spectra of DNA components can be used to predict the relative amounts of the bases within more complex molecules either by univariate analysis (i.e., by relating the base molecular ions to the sugar fragment ions within the nucleotides) or by multivariate analysis (e.g., principal component analysis). Our preliminary examination of four oligonucleotides shows promising results in that we can distinguish between two oligomers of similar composition using univariate and multivariate analysis, although additional studies are needed to expand this method to more complex oligomers.

Base Sequence↗
PubMed1987

Guanosine binding required for cyclization of the self-splicing intervening sequence ribonucleic acid from Tetrahymena thermophila.

We have converted the intramolecular cyclization reaction of the self-splicing intervening sequence (IVS) ribonucleic acid (RNA) from Tetrahymena thermophila into an intermolecular guanosine addition reaction. This was accomplished by selectively removing the 3'-terminal nucleotide by oxidation and beta-elimination; the beta-eliminated IVS thereby is no longer capable of reacting with itself. However, under cyclization conditions, a free guanosine molecule can make a nucleophilic attack at the normal cyclization site. We have used this guanosine addition reaction as a model system for a Michaelis-Menten kinetic analysis of the guanosine binding site involved in cyclization. The results indicate that functional groups on the guanine that are used in a G-C Watson-Crick base pair are important for the cyclization reaction. This is the same result that was obtained for the guanosine binding site involved in splicing [Bass, B. L., & Cech, T. R. (1984) Nature (London) 308, 820-826]. Unlike splicing, however, certain additional nucleotides 5' to the guanosine moiety make significant binding contributions. We conclude that the guanosine binding site in cyclization is similar to, but not identical with, the guanosine binding site in splicing. The same binding interactions used in cyclization could help align the 3' splice site of the rRNA precursor for exon ligation. We also report that the phosphodiester bond at the cyclization site is susceptible to a pH-dependent hydrolysis reaction; the phosphodiester bond is somehow activated toward attack by the 3'hydroxyl of a guanosine molecule or by a hydroxyl ion.

Animals↗