Unit 2: #2 Genetics: DNA Replication

What is the replication of DNA considered?

Replication is semiconservative

• Determined by Meselson and Stahl (1958)

What is the semiconservative model?

• Two strands of parental DNA separate 

• Each strand is template for synthesis of a new complementary strand 

• New strand is attached to old strand so half is always from the original 

What are the three proteins involved in replication?

• Before replication begins, the two strands of DNA must be unraveled and kept separate 

  • DNA helicase

    • Unwinds the double helix by breaking the hydrogen bonds between the base pairs 

  • Single stranded binding proteins (SSBs) 

    • Keeps the two strands apart, blocks H-bonding 

  • Topoisomerases 

    • Enzymes that relieves tension from DNA unwinding

    • Cuts both strands of DNA, then reseals cut strands 

Process of DNA Replication

• Replication begins at special sites called origins of replication 

• Two strands open up a replication “bubble” 

• Replication proceeds in both directions from bubble 

• Multiple replication bubbles form and fuse into each other 

• At each end of the replication bubble is a replication fork

Replication Fork

• *Y-shaped region where new strands of DNA elongating, where two strands of “old” DNA are still attached 

• Replication proceeds towards the fork on one strand and away from the fork on the other strand 

How are the complimentary strands built?

• DNA Polymerases 

  • Enzymes function in DNA replication and repair 

• DNA Polymerase III

  • Enzyme builds the complementary strand using template strand as a guide 

  • Adds complementary strand in 5’ to 3’ direction 

  • Needs an RNA primer as a starting point 

RNA Primers

• Made by enzyme primase 

• 10-60 nucleotides attached to template strand temporarily 

• Allows DNA Polymerase to begin elongation by adding free deoxyribonucleoside triphosphates to complementary strand 

Deoxyribonucleoside Triphosphates

• Similar to ATP 

• Deoxyribose instead of ribose sugar 

• Located in nucleus 

• Used by DNA Polymerase III to build complementary strand 

• Two phosphates removed and remaining nucleotide added to elongating strand

Problem with Antiparallel DNA strands

• DNA polymerase III adds nucleotides to the free 3’ end of growing strand (5’ → 3’ direction) 

• The two strands run antiparallel 

• Only one strand can be built continuously

What are the two strands found in DNA?

Leading & Lagging Strand

Leading Strand

• Uses the 3’ to 5’ template strand as its’ guide 

• Formed toward replication fork 

• Only one RNA primer required

Lagging Strand

• Made discontinuously in short fragments called Okazaki fragments

• Formed away from replication fork 

• Requires an RNA Primer for each fragment 

DNA Polymerase I

• Removes all RNA Primers and replaces them with correct deoxyribonucleotides 

• Enzymes DNA Ligase joins Okazaki fragments into one strand by forming phosphodiester bonds 

• As new strands form attached to “old” DNA, each pair twists into helix

Proofreading

• DNA polymerase I,II and III act as proofreaders of newly made strands

• DNA Polymerase II proofreads the DNA between rounds of replication as well 

• When mistake is detected, an exonuclease cuts out error

Solenoid

• A group of 6 nucleosomes coiled into cylindrical fibers 

• Further reducing the volume of DNA

• Solenoids act as strong storage units for non-coding sequences of DNA to protect it from potential damage

• DNA during interphase in form of nucleosomes or solenoids