DNA, Chromosomes and DNA Replication Dr.Aida Fadhel Biawi

DNA REPLICATION DNA replication is a biological process that occurs in all living organisms and copies their DNA; it is the basis for biological inheritance. The process starts when one double-stranded DNA molecule produces two identical copies of the molecule.

How does DNA replicate? DNA Replication is a semiconservative process that results in a double-stranded molecule that synthesizes to produce two new double stranded molecules such that each original single strand is paired with one newly made single strand.

Semiconservative replication would produce two copies that each contained one of the original strands and one new strand.

replication begins at specific sites on DNA molecule called "origins of replication, " origins are specific sequence of bases

mammalian DNA have many origins

The replication fork is a structure that forms within the nucleus during DNA replication. It is created by helicases, which break the hydrogen bonds holding the two DNA strands together.

Replication fork is where the parental DNA strands hasn't untwist Replication fork is where the parental DNA strands hasn't untwist. Replication bubbles allow DNA replication to speed up therefore the untwisted DNA would not be attacked by enzymes while replicating. . ( Which enzymes can attack DNA?? )

Specific enzymes & proteins recognize origins & bind DNA : 1- primase and DNA polymerase will find these specific portions and will bind to the template DNA at the correct location . ( DNA replication requires a RNA primer , primer synthesized by the enzyme primase , primer is a short strand RNA about 5 bases and RNA primer is complementary to DNA )

- new DNA synthesized by DNA polymerase , DNA polymerase binds to parent DNA strand with primer. - DNA polymerase sequentially adds deoxyribonucleotides to RNA primer , deoxyribonucleotides added have bases complementary to parent strand DNA . - The rate nucleotide additions in bacteria add about 500 bases/second while in mammels add about 50 bases/second ??!!

2-replication requires strand separation strand separation begins at origin of replication (Helicase) specific proteins prevent the two separated DNA strands from coming back together (single strand binding protein)

At origin of replication, one strand of DNA is made in a continuous manner (the leading strand) and the other in a discontinuous manner (the lagging strand(. DNA is made in only the 5-prime to 3-prime direction and the replication bubble opens the original double stranded DNA to expose both a 3-prime to 5-prime template (Leading strand template) and it complement. The lagging strand must be synthesized as a series of discontinuous segments of DNA. ?? These small fragments are called Okazaki fragments and they are joined together by an enzyme known as DNA ligase.

Function in DNA replication Single-Strand Binding (SSB) Proteins Enzyme Also known as helix destabilizing enzyme. Unwinds the DNA double helix at the Replication Fork. DNA Helicase Builds a new duplex DNA strand by adding nucleotides in the 5' to 3' direction. Also performs proof-reading and error correction. DNA Polymerase Bind to ssDNA and prevent the DNA double helix from re-annealing after DNA helicase unwinds it thus maintaining the strand separation. Single-Strand Binding (SSB) Proteins Relaxes the DNA from its super-coiled nature. Topoisomerase Re-anneals the semi-conservative strands and joins Okazaki Fragments of the lagging strand. DNA Ligase Provides a starting point of RNA (or DNA) for DNA polymerase to begin synthesis of the new DNA strand. Primase

DNA CONDENSATION A cell's genetic information, in the form of DNA, is stored in the nucleus. The space inside the nucleus is limited and has to contain billions of nucleotides that compose the cell's DNA. Therefore, the DNA has to be highly organized or condensed. There are several levels to the DNA packaging.

At the finest level, the nucleotides are organized in the form of linear strands of double helices. The DNA strand is wrapped around histones, a form of DNA binding proteins. Each unit of DNA wrapped around a histone molecule is called a nucleosome . The nucleosomes are linked together by the long strand of DNA. Characteristics of Histone Proteins - Octamere structure - Responsible for packaging DNA into nucleosomes - 4 different types: H2A, H2B, H3, H4

Nucleosome Structures Histone octamer 2 H2A 2 H2B 2 H3 2 H4

Beads on a String—10 nm Fiber

histones DNA protein purification H1 H3 H2A (= 1g per g DNA) H2B H4 Basic (arg, lys); + charges bind to - phosphates on DNA 09/01/12 22 SBL201 22

To further condense the DNA material, nucleosomes are compacted together to form chromatin fibers. The chromatin fibers then fold together into large looped domain. During the mitotic cycle, the looped domains are organized into distinct structures called the chromosomes. Packing of DNA into Chromatin

- histone-DNA complexes, referred to as nucleosomes, which are further folded into higher-order chromatin structures .

Nucleofilament or solenoid

30 nm fiber

Fig. 9 Orders of chromatin structure from naked DNA to chromatin to fully condensed chromosomes...

- Chromosomes are also used as a way of referring to the genetic basis of an organism as either diploid or haploid. Many eukaryotic cells have two sets of the chromosomes and are called diploid. Other cells that only contain one set of the chromosomes are called haploid.

A diploid cell has two sets of each of its chromosomes A human has 46 chromosomes (2n = 46) In a cell in which DNA synthesis has occurred all the chromosomes are duplicated and thus each consists of two identical sister chromatids Maternal set of chromosomes (n = 3) Paternal set of 2n = 6 Two sister chromatids of one replicated chromosome Two nonsister chromatids in a homologous pair Pair of homologous chromosomes (one from each set) Centromere