DNA Structure & Chromosomal Organisation PDF

1/26/2025 Week 7 DNA Structure & Chromosomal Organisation based on eBook chapter 7 DNA Structure https://www.youtube.com/watch?v=qy8dk5iS1f0  1/26/2025 DNA discovery – humble beginnings...

1/26/2025 Week 7 DNA Structure & Chromosomal Organisation based on eBook chapter 7 DNA Structure https://www.youtube.com/watch?v=qy8dk5iS1f0  1/26/2025 DNA discovery – humble beginnings Nuclei were first purified from pus cells in the 1860’s Friedrich Miescher First organelle ever purified DNA discovery – humble beginnings Purified the nuclei Chemical analysis revealed Hydrogen, Carbon, Oxygen, Nitrogen and Phosphorous This same substance was found in all cells he examined - Nuclein  1/26/2025 Chromosomes carry Genetic Information Early 1900’s – chromosomes carry the genetic information Chromosomes contain an acidic portion (DNA) and a basic portion (protein) DNA – 4 different nucleotides Proteins – 20 different amino acids Which was the important bit? DNA Protein Pneumonia answered the question Two strains of Streptococcus pneumoniae Strain S – capsule allowed infection Strain R – no capsule, no infection Frederick Griffith injected mice proving infection Mice injected with live R strain Mice injected with live S strain Mice live. No live R-strain cells in Mice die. Live S-strain cells in their their blood blood  1/26/2025 Pneumonia answered the question Frederick Griffith tried different experiments Mice were injected with heat-killed S-strain Mice injected with heat-killed S-strain Mice live. No live S-strain cells in their blood Pneumonia answered the question Mice injected with heat-killed S strain, plus live R-strain Mice injected with heat-killed S-strain, plus live R-strain Mice die. Live S-strain cells in their blood  1/26/2025 Transformation R-strain cells could now make capsules and infect Genetic information had passed from the dead S-strain to the live R strain - Transformation Nucleic Acid carries the Genetic Material 1944 Avery, MacLeod & McCarty proved nucleic acids, not proteins were the genetic material Again experimented with Pneumonia  1/26/2025 Nucleic Acid carries the Genetic Material Purified the components of S-strain cells Carbohydrates, fats, proteins and nucleic acids Each was mixed with live R-strain cells and injected into mice Live R-strain plus Carbohydrates Fats Proteins Nucleic Acid Live Live Live Died DNA not RNA carries the Genetic Material Purified nucleic acid was treated with either Rnase (destroys RNA) or Dnase (destroys DNA) DNA and RNA were added separately to the R-strain of S. pneumoniae Only the bacteria with the DNA added developed into S- strain DNA carries the genetic information DNA controls the synthesis of specific products  1/26/2025 DNA carries the Genetic Information Viruses to the rescue Bacteriophages finally proved DNA’s importance – 1950’s They are viruses that infect bacteria They are made up of only protein and DNA Perfect for determining which carries the genetic information  1/26/2025 Bacteriophage Contractile Tail fibres Hershey and Chase Experiments Used two radioactive compounds 35S used to label protein coats 32P used to label DNA Infected two different cultures of bacteria and figured out where the radioactivity was  1/26/2025 Hershey and Chase Experiments Bacterial Cell Radioactive protein Most 35S found in capsule (35S) supernatant DNA Labelled Phages infect Blender separates Phages Cells and Phages are bacteria outside the bacteria from separated by centrifugation cells and their components Bacterial Cell Protein capsule Most 32P found in pellet Radioactive DNA 32P Hershey and Chase Experiments Summary DNA gets injected into bacteria, protein remains outside DNA directs synthesis of new viruses DNA carries the genetic information  1/26/2025 Nucleic Acid Structures Two different types DNA (deoxyribonucleic acid) RNA (ribonucleic acid) Both are made up of nucleotide subunits Phosphate group Sugar (deoxyribose or ribose) Nitrogen-containing base Purines (A & G) or Pyrimidines (C & T/U) DNA RNA Nucleotide Components - Bases Purine short name, big structure Double ring Adenine & Guanine Adenine (A) Thymine (T) Uracil (U) Purine (in DNA) (in RNA) Pyrimidine Pyrimidine Pyrimidine long name, small structure Single ring Cytosine, Thymine & Uracil Guanine (G) Cytosine (C) Purine Pyrimidine  1/26/2025 Nucleotide Components – Sugars and Phosphate Sugars Deoxyribose Ribose (in DNA) (in RNA) Phosphate DNA vs RNA Deoxyribose Ribose sugar sugar Thymine base Uracil base Double- Single- stranded stranded  1/26/2025 DNA Nucleotides Adenine (A) Thymine (T) Phosphate group Sugar (deoxyribose) Guanine (G) Cytosine (C) Nucleotides join to form Polynucleotides A Nucleotides are joined to form a chain Polynucleotides have a direction G 5’ end - phosphate 3’ end – sugar C This sequence is 5’-AGCT-3’ T  1/26/2025 Chargaff’s rules of DNA composition Edwin Chargaff examined the DNA of numerous species in the 1940’s and discovered: 1. The number of Adenine residues always equals the number of Thymine residues A=T 2. The number of Cytosine residues always equals the number of Guanine residues C=G 3. The number of Purines always equals the number of Pyrimidines (A+G) = (C+T) Rosalind Franklin Used X-ray crystallography to study the physical structure of DNA Concluded: DNA is a helix Phosphates are on the outside Constant diameter  1/26/2025 Watson and Crick In 1953 described the model for DNA Used information from research available at the time Chagraff’s chemical nature of DNA Franklin’s physical structure of DNA Built models out of cardboard and wire Eventually made one to satisfy all the rules Watson and Crick DNA has two chains Sugars and Phosphates on the outside Chains are antiparallel Chains coil to form a helix Bases are paired across the chains via hydrogen bonds Pairing is specific A-T, C-G Watson and Crick and their Strands are complementary wire model  1/26/2025 DNA is a double-stranded helix DNA strands are antiparallel Reads Reads  1/26/2025 Watson & Crick’s Model Genetic information is stored in the sequence of bases in the DNA The model offers a molecular explanation for mutation The complementary sequence of bases explains how DNA is copied before each cell division DNA allows complexity At any point in the sequence there can be any one of 4 bases - A, C, G or T For a string of 3 nucleotides there are 64 (43) possible combinations (AAA, AAC, AAG, AAT, ACA, ACC…) For a string of n nucleotides there are 4n 10 nucleotides = 410 = 1,048,576 possibilities Human genome ~3.2 billion bases!  1/26/2025 An explanation for mutation Because DNA is a linear string of bases, any change in the order or number of bases can lead to an altered phenotype But what is RNA? A nucleic acid Single stranded A, C, G and U (Uracil) Ribose instead of deoxyribose Transfers genetic information from the nucleus to the cytoplasm The message for protein synthesis Also a component of ribosomes  1/26/2025 DNA vs RNA Three main types of RNA Messenger RNA (mRNA) Carries information from the DNA to the cytoplasm for protein synthesis Transfer RNA (tRNA) Transports amino acids to the ribosome Ribosomal RNA (rRNA) Combines with proteins to make the ribosome  1/26/2025 Three main types of RNA DNA’s Semi-conservative Replication One old strand is conserved in each new DNA molecule and one new one synthesized Each complementary strand of the DNA molecule is used as a template Proceeds in a 5’  3’ direction Occurs during the S phase of the cell cycle  1/26/2025 DNA Replication – Semi-conservative Semi-conservative One old strand and one new strand in each daughter DNA molecule Conservative Two DNA molecules generated, but one entirely new and one entirely old Dispersive Two DNA molecules generated, bits of old and bits of new in each DNA Replication Begins at origins of replication Hydrogen bonds are broken by helicase Primase forms a starting point DNA Polymerase III “reads” the sequence and joins on complementary bases DNA Polymerase III also does checks New DNA is made continuously on one strand, but in Okazaki fragments on the other DNA ligase fills in the gaps Proteins wind the strands into a helix  1/26/2025 DNA Replication – 4 Major Enzymes Helicase Finds the origin of replication Unzips the DNA by breaking hydrogen bonds, creating two single strands that can be used as templates  1/26/2025 Primase Creates a short RNA primer to act as starting point for DNA replication Provides a free OH group for DNA Polymerase III to add bases to Primase DNA Polymerase III DNA polymerase III synthesizes the new strand 5’-3’ It binds to the 3’ end of the RNA primer, reads the base on the single strand and pulls a complementary base towards it, joining it with a hydrogen bond Uses excess phosphates to provide the energy required  1/26/2025 DNA replication – Leading and Lagging Two new strands are formed One is continuous – leading strand One is formed from short fragments known as Okazaki fragments – lagging strand Okazaki Fragments  1/26/2025 DNA Ligase DNA Polymerase I converts RNA primers to DNA DNA Ligase joins the Okazaki fragments together to produce one long strand DNA Polymerase III Quality Control DNA Polymerase III also checks and corrects errors as it goes  1/26/2025 Bi-directional DNA replication Two replication forks are created from each origin of replication DNA synthesis occurs in both directions There is a leading strand and a lagging strand in both directions Replication fork Replication fork The Organisation of DNA into Chromosomes Over 2m of DNA in the nucleus Each chromosome consists of highly coiled double- stranded DNA DNA winds around histones to form nucleosomes 8 histone molecules join to form the histone core  1/26/2025 The Organisation of DNA into Chromosomes Nucleosomes condense DNA 6-7 times But DNA is compacted 5,000-10,000 times during replication! Compaction of DNA into Chromosomes Several models have “nucleosome” been proposed Chromatin (DNA + proteins) folds into loops that attach to a protein scaffold  1/26/2025 Centromeres and Telomeres Centromere Region where sister chromatids attach Telomere Capping on the ends of chromosome arms Role of Centromeres Different location on each chromosome Kinetochore forms at the centromere during cell division - important for attachment of spindle fibres Contain long stretches of repetitive DNA  1/26/2025 Role of Telomeres TTAGGG repeated hundreds of times Provide a structure to maintain DNA stability Not every repeat gets copied during replication Chromosomes get progressively shorter Rate limiting step in determining the number of times a cell can divide 50-60 divisions Role of Telomeres Telomerase protects telomeres during meiosis Binds to the end of the DNA and provides a short stretch of dsDNA Acts as a starting point for DNA Polymerase III to add more DNA  1/26/2025 Nuclear Organisation Each chromosomes has a designated region in the nucleus – chromosome territory Organisation is linked with function Flexible