DNA polymerase I

Summary

In this video, we explore the discovery and functions of DNA polymerase I, an enzyme first identified by Sir Arthur Kornberg in 1956. The enzyme was crucial in supporting Watson and Crick's hypothesis of DNA replication. DNA polymerase I performs three key activities: DNA polymerization from the 5' to 3' end, exonuclease activity from the 5' to 3' end, and proofreading through 3' to 5' exonuclease activity. These functions ensure accurate DNA replication by adding nucleotides, removing errors, and avoiding mutations in newly replicated DNA.

Takeaways

• 🧬 DNA Polymerase I was the first DNA polymerase discovered by Sir Arthur Kornberg in 1956.
• 🔬 The discovery was based on Watson and Crick's hypothesis that an enzyme in the cell could replicate DNA.
• 🧪 Kornberg conducted experiments using DNA samples, labeled dNTPs, and E. coli cell extract to test the hypothesis.
• 📈 The increase in counts per minute in the DNA confirmed the presence of an enzyme that adds dNTPs to DNA.
• 🧹 DNA Polymerase I was purified through various chromatography techniques.
• ⚙️ DNA Polymerase I has three major activities: 5' to 3' DNA polymerization, 5' to 3' exonuclease activity, and 3' to 5' proofreading.
• 🔄 5' to 3' DNA polymerization involves adding complementary dNTPs to the template DNA in the presence of magnesium ions.
• 🧩 The enzyme requires a primer for the polymerization process to extend in the 5' to 3' direction.
• ✂️ 5' to 3' exonuclease activity removes nucleotides from the DNA in the presence of single-stranded cuts or nicks.
• 🔍 3' to 5' exonuclease activity helps in proofreading by removing wrongly added nucleotides, preventing mutations in the newly replicated DNA.

Q & A

Who discovered DNA polymerase one and in what year?

DNA polymerase one was discovered by Sir Arthur Kornberg in 1956.

What hypothesis did Watson and Crick propose in 1953?

Watson and Crick proposed that there might be an enzyme in the cell that could replicate DNA.

How did Sir Arthur Kornberg confirm the presence of DNA polymerase one?

Kornberg confirmed the presence of DNA polymerase one by conducting experiments where he added E. coli cell extract to a DNA sample with labeled dNTPs. The incorporation of labeled dNTPs into the DNA indicated that an enzyme in the cell was replicating the DNA.

What are the three major activities of DNA polymerase one?

The three major activities of DNA polymerase one are: 1) 5' to 3' DNA polymerization activity, 2) 5' to 3' exonuclease activity, and 3) 3' to 5' proofreading activity.

What is required for the 5' to 3' DNA polymerization activity of DNA polymerase one?

The 5' to 3' DNA polymerization activity requires a primer that can be extended in the 5' to 3' direction, along with the presence of magnesium ions.

What role do magnesium ions play in DNA polymerization?

Magnesium ions help the oxyanion formed at the 3' end hydroxyl group to react with the alpha phosphoryl group of dNTPs, facilitating the DNA polymerization process.

What is the significance of the 5' to 3' exonuclease activity in DNA polymerase one?

The 5' to 3' exonuclease activity allows the polymerase to remove nucleotides in the 5' to 3' direction, which is important for repairing small single-stranded cuts on the DNA, also known as nicks.

What is 'nick translation' in the context of DNA polymerase one?

Nick translation refers to the simultaneous occurrence of the 5' to 3' exonuclease activity and the 5' to 3' polymerization activity, where the polymerase removes nucleotides at nicks and simultaneously adds new ones.

How does DNA polymerase one prevent mutations during DNA replication?

DNA polymerase one prevents mutations through its 3' to 5' exonuclease activity, which removes wrongly added nucleotides, ensuring the accuracy of DNA replication.

Why is the 3' to 5' exonuclease activity referred to as proofreading?

The 3' to 5' exonuclease activity is referred to as proofreading because it corrects errors by removing incorrectly added nucleotides during DNA replication, thereby maintaining the integrity of the newly synthesized DNA.