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The strength of the template effect attracting nucleotides to naked DNA.

Kervio E, Claasen B, Steiner UE, Richert C - Nucleic Acids Res. (2014)

Bottom Line: Combined with rate constants for the chemical step of extension and hydrolytic inactivation, our quantitative theory explains why some enzyme-free copying reactions are incomplete while others are not.For example, for GMP binding to ribonucleic acid, inhibition is a significant factor in low-yielding reactions, whereas for amino-terminal DNA hydrolysis of monomers is critical.Our results thus provide a quantitative basis for enzyme-free copying.

View Article: PubMed Central - PubMed

Affiliation: Institute for Organic Chemistry, University of Stuttgart, 70569 Stuttgart, Germany.

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Occupancy of extension site by the deoxynucleotide complementary to the templating base at 20°C, as calculated for different concentrations of 2′-deoxynucleotides 1a–t using binding constants reported in Table 1 or Table 2. Binding to (a) hairpins 2a, 2c, 2g or 4t, (b) template–primer duplexes 8tnt:9a–t and (c) template–primer duplexes 8tnt:9a–t in the presence of downstream-binding oligonucleotide 10a–t. Note the different scales of the x-axes in (a) and (b)/(c).
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Figure 8: Occupancy of extension site by the deoxynucleotide complementary to the templating base at 20°C, as calculated for different concentrations of 2′-deoxynucleotides 1a–t using binding constants reported in Table 1 or Table 2. Binding to (a) hairpins 2a, 2c, 2g or 4t, (b) template–primer duplexes 8tnt:9a–t and (c) template–primer duplexes 8tnt:9a–t in the presence of downstream-binding oligonucleotide 10a–t. Note the different scales of the x-axes in (a) and (b)/(c).

Mentions: With the binding constants in hand, we asked to what extent the reaction site of primer extension was occupied by the cognate nucleotide at a given concentration. Figure 8 shows calculated occupancies for different nucleotides and binding scenarios. When the reaction site is occupied, the primer terminus is protected from side reactions (38). An extension site occupied by the correctly paired monomer is also blocked from untemplated misincorporations. Untemplated reactions are common, and over-extension of primers, beyond the length of the template, are frequently observed (37,44). Finally, in the bound state, the activated nucleotide will be at least partially protected from side reactions with other nucleotides (unspecific polymerization, pyrophosphate formation, etc.) (61) and, being sterically less accessible, less prone to hydrolyze.


The strength of the template effect attracting nucleotides to naked DNA.

Kervio E, Claasen B, Steiner UE, Richert C - Nucleic Acids Res. (2014)

Occupancy of extension site by the deoxynucleotide complementary to the templating base at 20°C, as calculated for different concentrations of 2′-deoxynucleotides 1a–t using binding constants reported in Table 1 or Table 2. Binding to (a) hairpins 2a, 2c, 2g or 4t, (b) template–primer duplexes 8tnt:9a–t and (c) template–primer duplexes 8tnt:9a–t in the presence of downstream-binding oligonucleotide 10a–t. Note the different scales of the x-axes in (a) and (b)/(c).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4066754&req=5

Figure 8: Occupancy of extension site by the deoxynucleotide complementary to the templating base at 20°C, as calculated for different concentrations of 2′-deoxynucleotides 1a–t using binding constants reported in Table 1 or Table 2. Binding to (a) hairpins 2a, 2c, 2g or 4t, (b) template–primer duplexes 8tnt:9a–t and (c) template–primer duplexes 8tnt:9a–t in the presence of downstream-binding oligonucleotide 10a–t. Note the different scales of the x-axes in (a) and (b)/(c).
Mentions: With the binding constants in hand, we asked to what extent the reaction site of primer extension was occupied by the cognate nucleotide at a given concentration. Figure 8 shows calculated occupancies for different nucleotides and binding scenarios. When the reaction site is occupied, the primer terminus is protected from side reactions (38). An extension site occupied by the correctly paired monomer is also blocked from untemplated misincorporations. Untemplated reactions are common, and over-extension of primers, beyond the length of the template, are frequently observed (37,44). Finally, in the bound state, the activated nucleotide will be at least partially protected from side reactions with other nucleotides (unspecific polymerization, pyrophosphate formation, etc.) (61) and, being sterically less accessible, less prone to hydrolyze.

Bottom Line: Combined with rate constants for the chemical step of extension and hydrolytic inactivation, our quantitative theory explains why some enzyme-free copying reactions are incomplete while others are not.For example, for GMP binding to ribonucleic acid, inhibition is a significant factor in low-yielding reactions, whereas for amino-terminal DNA hydrolysis of monomers is critical.Our results thus provide a quantitative basis for enzyme-free copying.

View Article: PubMed Central - PubMed

Affiliation: Institute for Organic Chemistry, University of Stuttgart, 70569 Stuttgart, Germany.

Show MeSH
Related in: MedlinePlus