Passage
DNA polymerization is one of the most conserved mechanisms of genome replication. Synthesis of a complete DNA strand requires a template, primers, a polymerase enzyme, and sufficient deoxyribonucleotide triphosphates (dNTPs) . The DNA polymerase enzyme binds consecutive base pairs on the template strand and extends the double helix by adding dNTPs to the primer. The amino acid residues in the active site of DNA polymerase form hydrogen bonds with Watson-Crick donors and acceptors on incoming DNA nucleotides to facilitate base pairing.The formation of the DNA double helix creates opposing changes in entropy and enthalpy. Favorable bonding interactions via hydrogen bonds during Watson-Crick base pairing results in negative enthalpy, and restricted rotation and flexibility of the DNA backbone generates negative entropy. Scientists hypothesize that hydrogen bonding between bases not only stabilizes the double helix but is also crucial for selective and efficient replication.Analogs that are similar in size and shape to naturally occurring bases can be used to determine the influence of hydrogen bonding on base pair selectivity. To mimic the structure of deoxythymidine triphosphate (dTTP) , researchers synthesized dNTP derivatives of difluorotoluene (dFTP) , a nonpolar analog that lacks Watson-Crick hydrogen bonding. Klenow fragment (KF) polymerase, which has 3′-5′ but not 5′-3′ exonuclease activity, was incubated with a mixture of DNA template, primers, and dNTPs, including dFTP derivatives. The efficiency of dFTP and natural dTTP nucleotide incorporation into a growing primer strand by KF is shown in Figure 1.
Figure 1 Template-specific selection of dFTP and dTTP by the KF enzyme
Adapted from Moran S, Ren RX, Kool ET. A thymidine triphosphate shape analog lacking Watson-Crick pairing ability is replicated with high sequence selectivity. Proc Natl Acad Sci USA. 1997;94(20) :10506-11.
-The Klenow fragment used in the experiment would be able to perform which of the following repair processes?

Question

Passage
DNA polymerization is one of the most conserved mechanisms of genome replication. Synthesis of a complete DNA strand requires a template, primers, a polymerase enzyme, and sufficient deoxyribonucleotide triphosphates (dNTPs) . The DNA polymerase enzyme binds consecutive base pairs on the template strand and extends the double helix by adding dNTPs to the primer. The amino acid residues in the active site of DNA polymerase form hydrogen bonds with Watson-Crick donors and acceptors on incoming DNA nucleotides to facilitate base pairing.The formation of the DNA double helix creates opposing changes in entropy and enthalpy. Favorable bonding interactions via hydrogen bonds during Watson-Crick base pairing results in negative enthalpy, and restricted rotation and flexibility of the DNA backbone generates negative entropy. Scientists hypothesize that hydrogen bonding between bases not only stabilizes the double helix but is also crucial for selective and efficient replication.Analogs that are similar in size and shape to naturally occurring bases can be used to determine the influence of hydrogen bonding on base pair selectivity. To mimic the structure of deoxythymidine triphosphate (dTTP) , researchers synthesized dNTP derivatives of difluorotoluene (dFTP) , a nonpolar analog that lacks Watson-Crick hydrogen bonding. Klenow fragment (KF) polymerase, which has 3′-5′ but not 5′-3′ exonuclease activity, was incubated with a mixture of DNA template, primers, and dNTPs, including dFTP derivatives. The efficiency of dFTP and natural dTTP nucleotide incorporation into a growing primer strand by KF is shown in Figure 1.

Figure 1 Template-specific selection of dFTP and dTTP by the KF enzyme
Adapted from Moran S, Ren RX, Kool ET. A thymidine triphosphate shape analog lacking Watson-Crick pairing ability is replicated with high sequence selectivity. Proc Natl Acad Sci USA. 1997;94(20) :10506-11.
-The Klenow fragment used in the experiment would be able to perform which of the following repair processes?

Quizplus · Accepted Answer

11ecba2d_0ea1_01ca_a3bf_63e0710eacdf_MD0008_00 DNA polymerases are capable of detecting and removing incorrect nucleotides at the ends of a growing DNA strand. This function, referred to as proofreading, prevents mutations or errors in DNA replication from accumulating. Normally, DNA polymerases are equipped with both 5′-3′ and 3′-5′ exonuclease activity that allows them to remove and replace incorrect nucleotides at either end of a DNA strand. However, the passage states that the Klenow fragment (KF) enzyme described in the experiment does not have 5′-3′ exonuclease activity. KF can only proofread DNA in the 3′-5′ direction on the template strand, so only errors at the 3′ end of the growing strand can be repaired. (Choice A) KF would not remove thymine dimers from the 5′ end of section 1 because it is deficient in 5′-3′ exonuclease activity. Nucleotide repair enzymes are required to excise thymine dimers. (Choice C) Mismatch repair enzymes, which have endonuclease rather than exonuclease activity, are required to correct mismatched nucleotides within a DNA strand. (Choice D) Damaged bases are removed after replication by base excision repair enzymes with endonuclease activity. Educational objective: DNA polymerase I proofreads DNA and normally has exonuclease activity in the 5′-3′ as well as the 3′-5′ direction that allows it to remove primers and damaged or incorrect bases at the ends of the strand. It cannot fix mistakes in the middle of a strand; instead, base excision repair and nucleotide excision enzymes have endonuclease activity to remove damaged bases and mismatched nucleotides from the middle of a DNA strand, respectively.

Passage DNA Polymerization Is One of the Most Conserved Mechanisms of of Genome