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Everyone starts with just one fertilized egg. By adulthood, that single cell has turned into about 37 trillion cells, many of which continue to divide, creating the same amount of fresh human cells every few months.
But these cells present a formidable challenge. An average dividing cell should have a full copy of 3.2. a billion DNA base pairing, about once every 24 hours. The cell’s replication machinery does an amazing job of this, copying genetic material at an unsteady pace of about 50 base pairs per second.
Still, it’s too late to replicate the whole thing. human genome. If the cell’s copying machinery were initiated simultaneously at the tips of each of the 46 chromosomes, the longest chromosome (#1 at 249 million base pairs) would be completed in about 2 months.
“Of course, the way cells get around this is that they start replicating in multiple places,” says structural biologist James Berger of the Johns Hopkins University School of Medicine in Baltimore. Article on DNA replication in eukaryotes in 2021 Annual Review of Biochemistry. Yeast cells have hundreds of possible so-called origins of replication, and animals such as mice and humans have tens of thousands of replication origins scattered throughout their genomes.
“But that is its own challenge,” says Berger. “It’s about where to start and how to time everything.” Without controlling precision, some DNA can be copied twice, wreaking havoc in the cell.
Tight control of the initiation of DNA replication is particularly important to avoid this havoc. Researchers are now moving toward a fuller understanding of the molecular restraints and balances that each origin has evolved to initiate just one copy of her DNA to generate exactly one complete new genome. We are taking a step forward.
do it right, do it fast
Bad things can happen if replication doesn’t start properly. In order for DNA to be copied, the DNA double helix must be opened, and the resulting single strands each act as a template for building a new second strand, making them susceptible to cleavage. Or the process may die. “We really want to solve replication quickly,” says John Diffley, a biochemist at the Francis Crick Institute in London. Problems during DNA replication can disrupt the genome, which is often a critical step in pathways to cancer.
Some genetic disorders are also caused by: Problems with DNA replication. For example, Meyer-Gorlin syndrome, which is short stature, small ears, and small or no patella, is caused by mutations in several genes that help initiate the DNA replication process.
A tightly coordinated dance involving dozens of proteins is required for the DNA-copy machinery to initiate replication at the appropriate point in the cell’s life cycle. Researchers have a pretty good understanding of which proteins do what because they’ve managed to get DNA replication to happen in cell-free biological mixtures in the lab. They mimicked the first critical steps in the initiation of replication. Uses Yeast Derived Protein—the same kinds used to make bread and beer—and they mimicked much of the entire replication process Use the human version of the replication proteinthat too.
Cells control the initiation of DNA replication in a two-step process. The overall goal of this process is to control the action of key enzymes called helicases that unwind her DNA double helix in preparation for copying it. In the first step, an inactive helicase is loaded onto the DNA origin, where replication begins. In the second step, a helicase is activated to unwind the DNA.
Ready (Helicase loaded)…
This process begins with a cluster of six proteins located at the origin. This cluster, called ORC, shaped like a double ring Berger’s team found a convenient notch that can be slid over the DNA strand.
In baker’s yeast, a favorite of scientists studying DNA replication, these initiation sites are easy to find. These initiation sites have specific 11- to 17-letter core DNA sequences rich in adenine and thymine chemical bases. Scientists have watched ORCs grab DNA and glide along. Scan origin sequence until a suitable location is found.
But in humans and other complex lifeforms, the place of origin isn’t so sharply delineated, and it’s not entirely clear what settles and grabs onto the ORC, says Alessandro, a structural biologist at the Crick Institute. Costa saysI have written About DNA replication initiation in 2022 Annual Review of Biochemistry. Replication seems likely to initiate where the genome, which is normally tightly wrapped around proteins called histones, becomes loose.
Once the ORC is anchored on the DNA, it attracts a second protein complex containing a helicase that eventually unwinds the DNA. Costa et al. used electron microscopy to elucidate how ORC occurs. First lure one helicase, then another. Helicases are also ring-shaped, each open to wrap around double-stranded DNA. Then the two helicases approach again, facing each other on the DNA strand, like her two beads on a thread.
At first it just sits there like a car with no gas in the tank. They have not yet been activated and for now the cells are doing business as usual.
Prepare (activate helicase)…
Things really kick in when a key molecule called a CDK waves the blue flag, kicking off a chemical step that attracts more proteins. One of them, a DNA polymerase that builds new DNA strands (what Costa calls a “typewriter”), attaches to each helicase. Others activate helicases and become able to expend energy along the DNA.
When this happens, the helicase changes shape, pushing one DNA strand and pulling the other. This strains the weak hydrogen bonds that hold the two strands together by the bases (As, Cs, Ts, Gs) that normally make up the rungs of the DNA ladder. Two threads are torn. Costa et al. observed how the two helicases joined. Untwisting the DNA between two peopleThey then observed how the helicase kept unbound bases stable and out of the way.
go!
Initially, both helicases are wrapped around both DNA strands, but they can’t go this far because they just bump into each other. But then they each change position and spit one DNA strand or the other out of the ring. Now separated, they can push past each other, and replication proceeds rapidly.
Each helicase drives a motor along its single strand in the opposite direction to the other. They leave their starting point behind, tearing apart hydrogen-bonded base pairs as they move. DNA polymerase is right behind and he copies the DNA letter when it is released from its partner.
The CDK’s second job is to stop any more helicases from jumping to the origin. Therefore, there is one start of replication per origin, ensuring an adequate copy of the genome. However, the copy does not start at each site at the same time. The entire process of DNA replication in human cells takes about 8 hours.
There are still many things to solve. First, the copied DNA is not a bare double helix. It wraps around histones and binds to many other proteins that are busy switching genes on and off. make RNA copies of genes. How do these huddled proteins influence each other and keep each other out of the way?
Beyond this fascinating and basic biology—an amazing process essential to all life on Earth—there are implications for diseases such as cancer. Scientists already know that replication defects can destabilize DNA, and that unstable, mutation-prone genomes may be an early hallmark of cancer development.and they investigate further Relationship between replication proteins and cancer.
“I think there is an opportunity for therapeutic intervention in these systems once we have enough insight into how they work and what they are like,” says Berger. .
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