Biology 1st Paper (Botany) · Classes 11-12 · Chapter 1 · 1.4

DNA Replication.

Before a cell divides, it has to copy all of its DNA. The copy must be fast, and nearly perfect. How? One double helix comes undone, and each open strand becomes the pattern for a new one.

1. One double helix, two copies

The process by which a parent DNA makes a similar DNA is DNA replication. One double helix becomes two double helices. It happens in the S phase of the cell cycle, which sits in interphase, before the cell divides.

2. Three hypotheses

In 1956 three hypotheses of replication were put forward: conservative, semiconservative and dispersive. They ask one question: where do the old strands end up in the two new DNAs?

Conservative: the parent strands stay together, unchanged, while an all-new DNA is built.
Semiconservative: each new DNA holds one parent strand and one new strand.
Dispersive: the strands break up, and old and new pieces mix to form two DNAs.

The word “semiconservative” was first used by Stent, in 1957. This is the hypothesis that proved true: Matthew Meselson and Franklin Stahl showed it in 1958 by experiment on the bacterium E. coli. Suekka proved the same method in human HeLa cells in 1960, and Simon in the alga Chlamydomonas in 1961.

Three hypotheses are easier to see than to recite. Pick one, copy the DNA for two generations, and follow the old strands.

Now drop each feature into its hypothesis.

3. In prokaryotes and eukaryotes

The DNA of a prokaryote is a circle with no ends. Replication can start anywhere. The forks then run in both directions and meet half way round, so the job ends quickly. Up to a million base pairs are added every minute.

The DNA of a eukaryote is long and thread-like, with two ends. The pace is slower, 500 to 5000 base pairs per minute. Replication does not start at an end; it starts at many places in the middle at once. In Drosophila there are 50000 such sites.

4. What it needs, and the replisome

Replication needs four things: a template; nucleotide triphosphates, that is dATP, dGTP, dTTP and dCTP; the energy that comes from those triphosphates; and the enzymes of the replisome.

The complex of enzymes and helper proteins is the replication complex, or replisome. Its main enzyme is DNA polymerase. The others are helicase, primase, SSBP, gyrase and topoisomerase.

Six replisome parts and their work:
Topoisomerase: relieves over-twisting.
Helicase: unwinds the helix.
DNA polymerase: builds the strand 5′→3′ and proofreads.
SSBP: stays bound to the single strands.
Ligase: joins the Okazaki fragments.
Primase: makes the RNA primer.

5. What happens at the fork

The point on the double helix where replication begins is the origin of replication, or ori. There helicase uses ATP energy to break the hydrogen bonds and open the double helix into two single helices.

As the helix opens, twist tries to pile up in the strand ahead. Gyrase does not let it gather; in eukaryotes topoisomerase does this work. SSBP keeps the two separated strands from joining again. Each single helix then serves as a template.

From each origin, two replication complexes move off in opposite directions. The Y-shaped structure that forms as the helix opens is the replication fork.

DNA polymerase cannot begin on its own. First primase copies part of the template and makes a short stretch of a few RNA bases. This is the primer, and its end carries a free 3′-OH group.

DNA polymerase III can add new nucleotides only at a 3′-OH end, so a new DNA strand always grows from 5′ to 3′. When a nucleoside triphosphate joins, its last two phosphates leave as pyrophosphate and energy is released. One phosphate stays, and the piece becomes a nucleotide. The 5′-phosphate of the incoming nucleotide bonds to the 3′-OH of the previous one.

The two template strands run in opposite directions, so the two new strands cannot grow the same way. The strand that grows continuously toward the fork is the leading strand. The one made in pieces is the lagging strand. Its pieces are Okazaki fragments, and each fragment needs its own primer.

Set the fork running. The two new strands grow at the same time, but not in the same way.

Later DNA polymerase I swaps the primer for DNA, leaving a small gap. Ligase seals the gaps between the Okazaki fragments and makes the strand continuous. Now tap the nine points of Figure 1.33. Template strands are in black, new strands in blue.

Now put the whole process in order. Helicase opening the helix and SSBP holding it open really happen almost together, but put helicase first here.

6. Catching errors, repair, and why it matters

When replication ends, the replisome comes off and moves away. Because it starts at many places at once, the whole helix turns into two double helices in a short time.

Now and then a wrong nucleotide slips into a new strand; this is a mismatch. In humans there is one such error per 1000 genes. DNA's proofreading system catches it and repairs it. For example, if C has paired with A, the C is removed and T is put in its place.

When repair fails, disease follows. Mismatch causes one kind of human colon cancer. If UV damage cannot be repaired, the skin disease Xeroderma pigmentosum and skin cancer result; questions sometimes spell it Xerodesma.

Last, why it matters. DNA replication is vital for cell division and for forming gametes. For traits to pass from one generation to the next, it has to happen.

7. Check yourself

Three questions in the exam's style. Start with the first.

Think a little before the second.

And the last one.

The whole note on eight cards.

Source: Biology 1st Paper (Botany), Classes 11-12 (NCTB), Chapter 1 “Cell and Its Structure”, section 1.4.