Polynucleotides. VI. Molecular properties and conformation of polyribouridylic acid

Richards, E. G.; Flessel, C. Peter; Fresco, Jacques R.

doi:10.1002/bip.360010504

Cited by 158 publications

(58 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Uracil was chosen as the nontitrating base since it is a poor stacker (Richards et al 1963;Turner and Bevilacqua 1993); in contrast, guanine self-associates (Kim et al 1991) and has an N7 pK a that would interfere with the titration (Izatt et al 1971;Saenger 1984). The microscopic pk a s of the three oligonucleotides were determined by pH titrations monitored by UV absorbance (Fig.…”

mentioning

confidence: 99%

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting

Moody¹,

Lecomte²,

Bevilacqua³

2005

RNA

117

View full text Add to dashboard Cite

Perturbation of pK a values can change the favored protonation states of the nucleobases at biological pH and thereby modulate the function of RNA and DNA molecules. In an effort to understand the driving forces for pK a shifting specific to nucleic acids, we developed a thermodynamic framework that relates proton binding to the nucleobases and the helix-coil transition. Key features that emerge from the treatment are a comprehensive description of all the actions of proton binding on RNA folding: acid and alkaline denaturation of the helix and pK a shifting in the folded state. Practical experimental approaches for measuring pK a s from thermal denaturation experiments are developed. Microscopic pk a values (where k a is the acid dissociation constant) for the unfolded state were determined directly by experiments on unstructured oligonucleotides, which led to a macroscopic pK a for the ensemble of unfolded states shifted toward neutrality. The formalism was then applied to pH-dependent UV melting data for model DNA oligonucleotides. Folded-state pk a values were in good agreement with the outcome of pH titrations, and the acid and alkaline denaturation regions were well described. The formalism developed here is similar to that of Draper and coworkers for Mg 2+ binding to RNA, except that the unfolded state is described explicitly owing to the presence of specific proton-binding sites on the bases. A principal conclusion is that it should be possible to attain large pK a shifts by designing RNA molecules that fold cooperatively.

show abstract

mentioning

confidence: 99%

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting

Moody¹,

Lecomte²,

Bevilacqua³

2005

RNA

117

View full text Add to dashboard Cite

show abstract

“…The chain length of these polymers may be estimated to be 350 residues for poly(dA) and 250 residues for [3H]poly(dT) by using expressions derived for the corresponding ribopolynucleotides polyadenylic acid and polyuridylic acid [20,21]. The specific activity of the [3H]poly(dT) was 8 …”

mentioning

confidence: 99%

Excision of Pyrimidine Dimers from Ultraviolet‐Irradiated DNA by Exonucleases from Mammalian Cells

Lindahl

1971

European Journal of Biochemistry

View full text Add to dashboard Cite

Two DNA specific exonucleases, DNase I11 and DNase IV, are present in mammalian cell nuclei. The activity of the purified enzymes with ultraviolet-irradiated polydeoxynucleotides and DNA as substrates has been investigated. DNase I11 shows a very poor ability to release pyrimidine dimers, although excision can be demonstrated by using high concentrations of enzyme. In contrast, DNase IV liberates h e r s in an effective fashion in vitro, and is a likely candidate to serve in this function also in vivo. The pyrimidine dimers are released by DNase IV as parts of oligonucleotides containing 5-8 residues. Neither of the two exonucleases is markedly inhibited by caffeine.When living cells are exposed to ultraviolet light, the main lesions produced are pyrimidine dimers in DNA, in particular dimers between adjacent thymine residues [l]. Such dimers are formed with similar efficiency in bacterial and mammalian cells [2], and both types of cells possess repair mechanisms that reduce the damaging effect of these structural defects in the DNA. One mechanism of repair depends on the excision of the pyrimidine dimers, followed by repair replication a t the single-stranded gaps thus produced in the DNA, and subsequent rejoining of the strand interruptions [3-71. While this type of DNA repair was originally discovered and characterized in microbial systems, the processes of pyrimidine dimer excision, repair replication, and joining of broken strands have now also been demonstrated to occur in mammalian cells as a consequence of radiation damage [8-lo]. Moreover, this mode of DNA repair is apparently of physiological sigdcance in man, as a loss of the ability to remove pyrimidine dimers from DNA seems to be the cause of the hereditary disease Xeroderma pigmentosum in which the patients are abnormally sensitive to ultraviolet light [ 111.It has been shown in bacteria that the excision of pyrimidine dimers from DNA requires two different enzymes : an endonuclease that specifically attacks ultraviolet-irradiated DNA and catalyzes the formaUnusual Abbreviations. Poly(dA), a homopolymer of deoxyadenylate ; poly(dT), a homopolymer of deoxythymidylate; poly(dA) * poly(dT), a double-stranded polymer composed of poly(dA) and poly(dT) ; poly[d(A-T)], an alternating double-stranded copolymer of deoxyadenylate and deoxythymidylate.All tritium-labeled polymers are labeled in the thymine residues.tion of a single-strand break close to the dimer, and an exonuclease that attacks at the strand interruption and has the property of being able to effectively release pyrimidine dimers from DNA [12,13]. The situation in mammalian cells again seems similar, as cells from a case of Xeroderma pigmentosum could excise damaged residues from irradiated DNA if adjacent single-strand breaks were introduced by non-enzymatic artificial means, i. e . the required endonuclease activity apparently was lacking but the exonuclease was present [la].Two different DNA-specific exonucleases, localized in the cell nuclei, have recently been found in mammalian tissues....

show abstract

“…First, as briefly noted above, binding of either DOPP to poly(U) induces an increase in the UV absorbance of poly(U) at 260 nm, with a small red shift of the maximum peak frequency (from 260 to ca. 265 nm) (18,19). Because neither 1 nor 2 shows any significant absorption in the 200-350-nm scanning range, these changes indicate that complexation of the protonated DOPPs to the poly(U) induces a conformational transition of the poly(U) in which any inherent stacking of the uridines has been disrupted [i.e., any inherent helical character of the poly(U) is converted into random coil as the poly(U) assumes a structure that is closer to its denatured form] (19,20).…”

Section: Resultsmentioning

confidence: 99%

Deoxypolypeptides bind and cleave RNA

Cheng

Mahendran

Gonzalez

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

We have prepared L-and D-deoxypolypeptides (DOPPs) by selective reduction of appropriately protected polyhistidines with borane, reducing the carbonyl groups to methylenes. The result is a chiral polyamine, not amide, with a mainly protonated backbone and chirally mounted imidazolylmethylene side chains that are mostly unprotonated at neutrality because of the nearby polycationic backbone. We found that, in contrast with the D-octahistidine DOPP, the L-octahistidine DOPP is able to cooperatively bind to a D-polyuridylic acid RNA; this is consistent with results of previous studies showing that, relative to D-histidine, L-histidine is able to more strongly bind to RNA. The L-DOPP was also a better catalyst for cleaving the RNA than the D-DOPP, consistent with evidence that the L-DOPP uses its imidazole groups for catalysis, in addition to the backbone cations, but the D-DOPP does not use the imidazoles. The L-DOPP bifunctional process probably forms a phosphorane intermediate. This is a mechanism we have proposed for models of ribonuclease cleavage and for the ribonuclease A enzyme itself, based on our studies of the cleavage and isomerization of UpU catalyzed by imidazole buffers as well as other relevant studies. This mechanism contrasts with earlier, generally accepted ribonuclease cleavage mechanisms where the proton donor coordinates with the oxygen of the leaving group as the 2-hydroxyl of ribose attacks the unprotonated phosphate. T he enzyme ribonuclease A (RNase A) hydrolyzes RNA with bifunctional catalysis, using His-12 as a base and protonated His-119 as an acid to form the first products, a 2,3-cyclic phosphate of one ribose and the free next ribose with a new terminal 5′-hydroxyl group (1, 2). Lys-41 is also involved in stabilizing intermediate anions (3). In a second step the cyclic phosphate is hydrolyzed using the now-protonated His-12 and now-deprotonated His-119 in essentially a reverse of the cyclization step, with water replacing the second ribose unit (4). In the classical mechanism of this process it had been generally accepted, including in textbooks, that the protonated His-119 donates its proton to the leaving group whereas the 2′-hydroxyl group attacks the phosphate (1, 2) [isotope studies show that two protons are "in flight" during the hydrolysis step, essentially the reverse of the cyclization, both moving at the same time (5)]. We also showed the two-proton-in-flight process in a model system (6).However, our extensive studies on the cleavage of RNA with imidazole-imidazolium catalysts and on catalysis by RNase A itself indicated that a different mechanism is used ( Fig. 1) (7, 8). The proton donor BH + hydrogen binds to a phosphate oxyanion and donates the proton to the phosphate as the ribose 2′-hydroxyl adds while losing a proton to base :B. This process results in an intermediate phosphorane species. This then expels the 5′-hydroxyl group of the second nucleotide to cleave the P-O bond in a second step. We proposed this two-step phosphorane process to be the preferred path for th...

show abstract

Polynucleotides. VI. Molecular properties and conformation of polyribouridylic acid

Abstract: Q The following abbreviations have been used: poly U = polyribouridylic acid; RNA = ribonucleic acid; DNA = deoxyribonucleic acid; poly A = polyriboadenylic acid; RNase = ribonuclease; UMP = uridinemonophosphate.

Cited by 158 publications

References 30 publications

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting

Excision of Pyrimidine Dimers from Ultraviolet‐Irradiated DNA by Exonucleases from Mammalian Cells

Deoxypolypeptides bind and cleave RNA

Contact Info

Product

Resources

About

Polynucleotides. VI. Molecular properties and conformation of polyribouridylic acid

Abstract: Q The following abbreviations have been used: poly U = polyribouridylic acid; RNA = ribonucleic acid; DNA = deoxyribonucleic acid; poly A = polyriboadenylic acid; RNase = ribonuclease; UMP = uridinemonophosphate.

Cited by 158 publications

References 30 publications

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pKa shifting

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pKa shifting

Excision of Pyrimidine Dimers from Ultraviolet‐Irradiated DNA by Exonucleases from Mammalian Cells

Deoxypolypeptides bind and cleave RNA

Contact Info

Product

Resources

About

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting

Linkage between proton binding and folding in RNA: A thermodynamic framework and its experimental application for investigating pK_a shifting