DNA Replication

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Leading Strand

Pear Tree = Parent strand 3-prime end; telephone in mirror = telomere; lasso protein bar = single-strand binding protein

Lagging Strand

primate painting rhino heads = RNA primase
construction crew member = lagging strand
"Okey-dokey" zoo keeper = Okazaki fragments
liger (lion/tiger) Pac-Man = DNA ligase

RNA Base Units

rhino heads = RNA base unit
dinosaur heads = DNA base unit; parrot Pac-Man = DNA polymerase
telephone in mirror = telomere

DNA Replication

DNA in all organisms is a double-strand helix made of nucleotides with two pairs of complementary bases. The double helix is zipped open and new nucleotides with bases complementary to the opened strands are added in a 5' to 3' direction, with different ways of formation of the two new daughter strands called leading and lagging strands. Though the process is similar in prokaryotes and eukaryotes, the enzymes used are different.

DNA

DNA is the genetic material that is found in the nucleus of all living organisms. It is found directly in the cytoplasm of unicellular prokaryotes. In eukaryotes, it is bound within the membranes of the nucleus, or in organelles like mitochondria and chloroplast, making it one of the main difference between these two kinds of organisms. The DNA in prokaryotes is a single loop, while it is long and unlooped in eukaryotes. The long DNA in eukaryotes is tightly coiled around proteins to give chromosomes.

 

DNADNA contains hereditary information that is used to synthesis proteins, determine the structure and functioning of a cell, as well as the growth and development of the organism.

Structure and Chemistry of DNA

The DNA is a double-stranded helix made of basic units called nucleotides. Each nucleotide is made of deoxyribose sugar, phosphate, and base. The deoxyribose sugar of one nucleotide is bonded with the phosphate of the next by strong bonds to make up the backbone of the strand, and the complementary bases make up the rungs of the double helix, bonded by hydrogen bonds which connect the two strands and create a strong DNA molecule. One strand ends in a 5 prime (5') at the phosphate end, and the other strand ends at the 3 prime (3') end at the deoxyribose end.

There are four bases in all DNA of every organism. The complementary pairs are adenine and thymine, and guanine and cytosine. The complementary nature of the bases ensures the complementary sequence of the two strands and is what makes replication of DNA possible. The order or sequence in which the bases occur is the important message that DNA transfer, and is called the genome. Before a cell divides, it needs to duplicate its DNA, so that each daughter cell has an exact DNA copy.

Replication

When DNA replicates, the two strands are separated and both are used as templates, so that the two new DNA have one old strand and one newly added strand. This is called semi-conservative replication, and is responsible for the stable transfer of genetic information. There are two main stages of replication.

  1. Replication Fork Formation

dna-forkThe DNA that is tightly coiled is unwound to reveal the long double helix DNA strand. Segments of the two strands are then separated or unzipped, with the help of the enzyme Helicase, which breaks the hydrogen bonds between the two complementary bases to form the y-shaped replication fork. The point at which the fork occurs is called the origin of replication(ORI). The ORI exposes the 3' and 5' ends of the two strands.

  1. Addition of New Nucleotides

 Nucleotides can be only added starting with a phosphate molecule or from the 5' end, complementary to the template that ends in 3'. This means that the process in which the two strands are formed differ. The old DNA strand ending in 3' forms the template for the leading strand, and the other strand ending in 5' is the template for the formation of the lagging strand.

DNA ReplicationA DNA polymerase enzyme attaches itself to the 3' or deoxyribose ended strand and adds nucleotides with bases complementary to bases on the old strand to form the leading strand from 5' to 3' in the direction moving towards the replication fork. This gives the first new double helix daughter DNA.

The lagging strand formation is more complicated, and happens in sections called Okazaki fragments. Many such segments maybe formed simultaneously. Initially, an RNA primer is attached by a Primase to a point on the 5' end strand, so that the first free nucleotide in the Okazaki fragment is again a 3' end. Then a second kind of DNA polymerase attaches itself and starts to add new complementary nucleotides in the opposite direction away from the replication fork. When a Okazaki fragment is complete, the RNA primer is removed and replaced by appropriate complementary nucleotides. Another DNA enzyme, Ligase, is used to bond the various fragments together to give a continuous strand that forms part of the second double helix daughter DNA.

Differences between DNA Replication in Prokaryotes and Eukaryotes

There are many differences between prokaryotes and eukaryotes. In prokaryotes, the DNA replication happens in the cytoplasm. Since the DNA is short, there is only one ORI, with one replication fork and one replication bubble. The enzymes used are simple proteins, and both strands are synthesised by DNA polymerase III. The replication is rapid, with 2000 base pairs being formed each second, with the aid of large Okazaki fragments of 1000-2000 nucleotides. The DNA replication is terminated by Topoisomerase II, which also separates the two interlocked loops of daughter DNAs.

 In eukaryotes, the DNA replication happens in the nucleus. Since DNA size is considerably larger, there are many ORIs, replication forks and replication bubbles. The enzymes used are multi-complex proteins, where the leading strand is formed by DNA polymerase epsilon (?), and the lagging strand by DNA polymerase delta (?). The speed of replication is slow, with 200 base pairs being formed per second, with small Okazaki fragments of 100-200 nucleotides. Telemerase is the enzyme used to terminate the replication.

DNA Replication

  1. Zoom: whole scene

Hot Spot: reptile in box = DNA replication; helicopter Pac-Man = DNA helicase; ice top-hat Pac-Man = DNA topoisomerase; parrot Pac-Man = DNA polymerase; lead race car = leading strand

Learning: The process of DNA replication, represented in this CoursePic as reptile in box, is complex and elegant. It begins when DNA helicase, shown as helicopter Pac-Man, unzips the double helix into what's called a replication fork. This exposes two strands ready for the addition of new nucleotides. Following the DNA helicase along the leading strand, pictured as lead race car, and regulating when necessary, is DNA topoisomerase, or ice top-hat Pac-Man. This activity readies the strand so the DNA polymerase enzyme can start adding nucleotides.

Story: The zoo is finally ready to begin building a reptile habitat in its popular Swamp Land attraction. The toothily grinning reptile in box is happy to help out by way of extending the family line. The local news outlet has dispatched helicopter Pac-Man to follow the construction and other activity. To rotect their investments, the wealthy patrons have hired ice top-hat Pac-Man as their stone-cold regulator. Hey, he's good at his job, and somebody's gotta keep an eye on self-described "DNA polymerase pro" parrot Pac-Man, right?

  1. Zoom: tree; telephone in mirror; lasso protein bar

Hot Spot: Pear Tree = Parent strand 3-prime end; telephone in mirror = telomere; lasso protein bar = single-strand binding protein

Learning: At the 3-prime end of the leading strand, symbolized by the tree, a telomere, seen here as telephone in mirror, serves as a cap to preserve interior elements. The 5-prime end of the chromosome is also capped. Single-strand binding proteins, represented by the lasso protein bars, act to prevent the entire replication fork from reforming.

Story: With a leafy tree for shade, the grinning 'gator is content to observe as lasso protein bar snaps a rope around the strand to keep it open as parrot Pac-Man to adds new nucleotides. Old poly is one hard-working cracker snapper! In fact, if not for the recently installed telomere, he'd probably run right off onto the lawn.

  1. Zoom: primate painting rhino heads; construction crew member; "Okey-dokey" zoo keeper; liger (lion/tiger) Pac-Man

Hot Spot: primate painting rhino heads = RNA primase; construction crew member = lagging strand; "Okey-dokey" zoo keeper = Okazaki fragments; liger (lion/tiger) Pac-Man = DNA ligase

Learning: With the DNA helix opened, the RNA primase, which is illustrated by primate painting rhino heads, moves along in the wake of the DNA helicase and synthesizes a primer to enable the actual replication. This uracil primer extends only about 10 nucleotides long before the DNA polymerase starts adding new material. Since the replication process is anti-parallel, nucleotides are added to the lagging strand and are pictured here as construction crew members in short DNA sections called Okazaki fragments, or the "Okey-dokey" zoo keeper. DNA polymerase can make segments, but cannot join them together. Another enzyme, DNA ligase, shown as liger Pac-Man, binds neighboring Okazaki fragments and finishes replication in the lagging strand.

Story: Watching the rough labor from his cool shade, reptile in box wonders at the display of energy, especially that of the primate painting rhino heads. The construction crew member looks on, knowing he got off easy thanks to his connections in lagging strand admin's 5-prime office. Even the zoo-keeper, old "Okey-dokey" Okazaki himself, doesn't seem to mind the snake draped across his shoulders.

  1. Zoom: rhino heads; dinosaur heads; parrot Pac-Man; telephone in mirror

Hot Spot: rhino heads = RNA base unit; dinosaur heads = DNA base unit; parrot Pac-Man = DNA polymerase; telephone in mirror = telomere

Learning: When the replication nears completion, and the base units of DNA and RNA, pictured respectively as dinosaur heads and rhino heads, have been variously paired by DNA polymerase, the telomerase enzyme binds to a specialized RNA molecule. Then, using it as a template, lengthens telomere's lagging strand by adding nucleotides. At this point, termination is reached.

Story: What a team! Swamp Land's reptile habitat is just about complete, and the entire sweating crew of parrot Pac-Man, rhino heads and dinosaur heads is approaching clock-out time, and  perhaps a quick call home on the telephone in mirror to see what's for dinner. 

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