Affinity of guanosine derivatives for polycytidylate revisited

Kanavarioti, Anastassia; Hurley, T. B.; Baird, Eldon E.

doi:10.1007/bf00170668

Cited by 11 publications

(49 citation statements)

References 27 publications

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“…Reactions were carried out in a 0.5 M HEPES aqueous buffer at pH 7.95 + 0.05 in the presence of 1.2 M NaC1 and 0.2 M MgC12 at 23 _+ 0.1°C; these are the same conditions used in Kanavarioti et al (1993Kanavarioti et al ( , 1995. Reactions were monitored for only a few hours so that no more than 15% of monomer was consumed, ensuring that the oligoguanylate products always remain the minor component of the system.…”

Section: Resultsmentioning

confidence: 99%

“…1 Kanavarioti et al (1995) d Pseudo-first-order rate constants determined as described in Kanavarioti et al (1993); see also Eqs. (2) and (3).…”

Section: Resultsmentioning

confidence: 99%

“…Poly(C) forms 1:1 complexes with guanosine derivatives at pH > 7 and 2:1 complexes at pH < 7 (Howard et al 1964;Davies and Davidson 1971). Evidence that under our conditions at pH 8, poly(C) forms a 1:1 complex with 2-MelmpG comes from the binding isotherm (Kanavarioti et al 1995) and from the experiments with [2-MeImpG] > or >> [poly(C)] ran to completion. For example, the yield of oligoguanylates with 0.002 M poly(C) and 0.02 M monomer is approximately 0.002 M, expressed in guanosine equivalents, whereas with 0.03 M template and 0.04 M monomer the yield is about 0.03 M. These high yields are only consistent with a 1:1 complex and not with a 2C: 1G complex, mainly because at the temperature of incubation the oligoguanylates do not dissociate from the template and there is no turnover of the reaction.…”

Section: Calculation Of Occupancy and Amount Of Template-bound Monomermentioning

confidence: 94%

“…(6), [M]tot is the total monomer concentration. Q = 180 M -I and q = 2.22 M -1 were determined from the binding isotherm at 23°C for [poly(C)] ranging from 1.78-7.55 mM (see Kanavarioti et al 1995). Assuming that the binding isotherm remains valid at poly(C) concentrations higher than 7.55 ms, the determined values of q and Q can be used to calculate Table 1.…”

Section: Calculation Of Occupancy and Amount Of Template-bound Monomermentioning

confidence: 99%

“…In an earlier report (Kanavarioti et al 1993), we showed the feasibility of a thorough kinetic study by determining rate constants for each step of the poly(C)-directed polymerization of 2-MelmpG (Inoue and Orgel 1982). We have also measured the binding of 2-MelmpG on poly(C) under the conditions of the polymerization reaction (Kanavarioti et al 1995).…”

Section: Introductionmentioning

confidence: 99%

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Faster rates with less catalyst in template-directed reactions

Kanavarioti

Baird

1995

J Mol Evol

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We have recently shown that the polycytidylic acid-directed polymerization of guanosine 5'-monophosphate 2-methylimidazolide (2-MelmpG) is amenable to kinetic study and that rate determinations as a function of 2-MelmpG concentration can reveal much mechanistic detail (Kanavarioti et al. 1993). Here we report kinetic data which show that, once the reaction has been initiated by the formation of dimers, the elongation of dimers to form longer oligomers is accelerated by decreasing polycytidylate (poly(C)) concentration from 0.05 to 0.002 M. This result is consistent with the previously proposed mechanism. The increase in the observed pseudo-first order rate constant for formation of the trimer, k3', and the corresponding constant for formation of oligomers longer than the trimer, k i' (k i' is independent of oligomer length for i I> 4), with decreasing template concentration for a given monomer concentration is attributed to an increase in template occupancy as template concentration is reduced.

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Section: Resultsmentioning

confidence: 99%

“…1 Kanavarioti et al (1995) d Pseudo-first-order rate constants determined as described in Kanavarioti et al (1993); see also Eqs. (2) and (3).…”

Section: Resultsmentioning

confidence: 99%

Section: Calculation Of Occupancy and Amount Of Template-bound Monomermentioning

confidence: 94%

Section: Calculation Of Occupancy and Amount Of Template-bound Monomermentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Faster rates with less catalyst in template-directed reactions

Kanavarioti

Baird

1995

J Mol Evol

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Preference for Internucleotide Linkages as a Function of the Number of Constituents in a Mixture

Kanavarioti

1998

J Mol Evol

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Phosphoimidazolide-activated ribomononucleotides (*pN; see Scheme I) are useful substrates for the nonenzymatic synthesis of oligonucleotides. In the presence of metal ions dilute neutral aqueous solutions of *pN (0.01 M) typically yield only small amounts of dimers and traces of oligomers; most of *pN hydrolyzes to yield nucleoside 5'-monophosphate (5'NMP). An earlier investigation of *pN reactions in highly concentrated aqueous solutions (up to 1.4 M) showed, as expected, that the percentage yield of the condensation products increases and the yield of the hydrolysis product correspondingly decreases with *pN concentration (Kanavarioti 1997). Here we report product distributions in reactions with one, two, or three reactive components at the same total nucleotide concentration. *pN used as substrates were the nucleoside 5'-phosphate 2-methylimidazolides, 2-MeImpN, with N = cytidine (C), uridine (U), or guanosine (G). Reactions were conducted as self-condensations, i. e., one nucleotide only, with two components in the three binary U,C, U,G, and C,G mixtures, and with three components in the ternary U,C, G mixture. The products are 5'NMP, 5',5'-pyrophosphate-, 2',5'-, 3', 5'-linked dimers, cyclic dimers, and a small percentage of longer oligomers. The surprising finding was that, under identical conditions, including the same total monomer concentration, the product distribution differs substantially from one reaction to another, most likely due to changing intermolecular interactions depending on the constituents. Even more unexpected was the observed trend according to which reactions of the U,C,G mixture produce the highest yield of internucleotide-linked dimers, whereas the self-condensations produce the least and the reactions with the binary mixtures produce yields that fall in between. What is remarkable is that the approximately two-fold increase in the percentage yield of internucleotide-linked dimers is not due to a concentration effect or a catalyst, but to the increased complexity of the system from a single to two and three components. These observations, perhaps, provide an example of how increased complexity in relatively simple chemical systems leads to organization of the material and consequently to chemical evolution. A possible link between prebiotic chemistry and the postulated RNA world is discussed.

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Effects of Monomer and Template Concentration on the Kinetics of Nonenzymatic Template-Directed Oligoguanylate Synthesis

1998

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To identify key parameters which influence the efficiency of nonenzymatic template-directed oligonucleotide reactions, a kinetic study of oligoguanylate synthesis on a polycytidylate (poly(C)) template has been performed. This oligomerization is satisfactorily described by three kinetic processes: (i) dimerization to form the first primer (k 2), (ii) elongation of a preformed primer (k i , i ≥ 3), and (iii) hydrolysis of the monomer to form deactivated material (k h), with k h < k 2 < k i . This is the first reported study that includes rate determinations of k i as a function of the concentration of both poly(C) template and the activated monomer, guanosine 5‘-monophosphate-2-methylimidazolide (2-MeImpG), in the range 2 mM ≤ [poly(C)] ≤ 50 mM and 5 mM ≤ [2-MeImpG] ≤ 50 mM. k i values determined under conditions where the template is fully saturated with monomer are practically independent of both monomer and polymer concentration and thus strongly support a template-directed elongation model. Values of k i determined with a partially occupied template support a mechanism wherein the reaction of the oligonucleotide primer with a template-bound monomer is assisted by the presence of two additional downstream template-bound 2-MeImpG molecules. Comparison between the kinetic parameters obtained here and the ones determined in the montmorillonite-catalyzed oligoadenylate polymerization allows the proposition that the ratio of the rate constants k i /k h determines efficiency and the ratio k i /k 2 determines the degree or extent of a polymerization. These conclusions provide new design principles for the optimization of nonenzymatic polymerizations.

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Affinity of guanosine derivatives for polycytidylate revisited

Cited by 11 publications

References 27 publications

Faster rates with less catalyst in template-directed reactions

Faster rates with less catalyst in template-directed reactions

Preference for Internucleotide Linkages as a Function of the Number of Constituents in a Mixture

Effects of Monomer and Template Concentration on the Kinetics of Nonenzymatic Template-Directed Oligoguanylate Synthesis

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