Principles of Biochemistry Volume 3 - A. Lehninger 1985

Molecular Mechanisms of Genetic Information Transfer
DNA Replication and Transcription
DNA polymerases can detect and correct errors

It has been established that the error rate in E. coli METABOLISM/36.html">DNA Replication does not exceed 1 in 10⁹-10¹⁰ NUCLEOTIDES. Since the E. coli chromosome contains approximately 4.5 ∙ 10⁶ Base Pairs, only a single incorrect nucleotide is incorporated per 10,000 Cells undergoing one division cycle. For a long time, it was believed that such a high degree of fidelity in reproducing Genetic information was determined entirely by the accuracy of Watson-Crick base pairing between the template and newly synthesized (daughter) strands. However, subsequent analysis revealed that if replication accuracy depended solely on correct base pairing, the error rate would be significantly higher—approximately 1 in 10⁴-10⁵ residues. Therefore, to account for such a low in vivo error rate, one must postulate the involvement of one or more additional factors in the replication process.

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Fig. 28-15. Proofreading via the 3'-exonuclease activity of DNA polymerase.

A more detailed study of The properties of highly purified DNA polymerases has provided at least a partial answer regarding The Nature of these factors. Recall that DNA polymerases I and III possess three distinct enzymatic activities. We have already seen how the enzyme Functions as a polymerase and how it can remove nucleotide residues from the 5'-end of a DNA fragment. However, the 3'-exonuclease activity of DNA polymerases I and III long puzzled researchers, as it implied that these Enzymes are capable of “backing up,” excising 3'-terminal nucleotides in the direction opposite to that in which they act as polymerases. The 3'-exonuclease activity of DNA polymerases I and III serves as a mechanism to inspect the newly synthesized DNA strand and correct errors made by the enzyme during its polymerization function. If DNA polymerase incorporates an incorrect nucleotide, the enzyme itself can recognize the inability of this nucleotide to form a correct pair with the corresponding template nucleotide (Fig. 28-15). In this case, the enzyme moves backward and excises the incorrect nucleotide from the 3'-end of the strand, after which the polymerase resumes incorporating correct nucleotides, i.e., continues its normal progression in the 5'→3' direction. Thus, as the Replication fork moves along the template, each incorporated nucleotide is inspected. The proofreading action of DNA polymerase is highly efficient; it increases replication fidelity by at least a factor of 10⁴. The overall error rate results from mistakes made by the enzyme during polymerization and during the proofreading correction process; it does not exceed one error per 10⁹-10¹⁰ nucleotide residues.

It is crucial to note that replication proceeds with a significantly higher degree of fidelity than Transcription and Translation Processes. Frequent replication errors would severely jeopardize species survival and viability. Errors in transcription and translation are far less dangerous, as they affect RNA or Protein Synthesis in only a single Cell and do not alter the subsequent Lineage of the species. Proofreading by DNA polymerase is likely only one of the pathways ensuring high replication fidelity. It is possible that the exceptionally complex Organization OF THE replication process and the involvement of numerous Proteins are required precisely to achieve this goal. Interestingly, some eukaryotic DNA polymerases lack proofreading activity. Apparently, eukaryotes ensure the precision and reliability of replication through alternative mechanisms.