FFP1_44 Genome Structure 2022-23 PDF

Royal College of Surgeons in Ireland – Medical University of Bahrain Genome structure Module : Foundations for Practice 1 Code : FFP1-44 Class : year 1 semester 1 Lecturer : Paul O’Farrell Date : 11 October 2022 Objectives 1. Outline the major steps of information processing in biology from...

Royal College of Surgeons in Ireland – Medical University of Bahrain Genome structure Module : Foundations for Practice 1 Code : FFP1-44 Class : year 1 semester 1 Lecturer : Paul O’Farrell Date : 11 October 2022 Objectives 1. Outline the major steps of information processing in biology from storage in the form of DNA through to generation of pure protein 2. Describe the basic structure of DNA, the nature of base pairing, and the types of bonds that hold DNA together 3. Describe the higher-order structuring of DNA (levels of folding) 4. Outline histone modification as a mechanism for regulating gene expression 5. Describe the fundamental elements of chromosomes and their organisation in the human genome 6. Describe the composition of human nuclear and mitochondrial genomes Genes, information and proteins Proteins determine the structure & biological activity of cells One ‘gene’ is a length of DNA that directs the synthesis of one protein* The full complement of DNA is called the ‘genome’ the human genome contains approximately 21,000 genes *There are also some genes that direct the synthesis of ‘functional’ RNA molecules, like tRNA. Information flow Transmission “The Central Dogma” of molecular biology - Francis Crick 1956 Deoxyribonucleic acid (DNA) A long thread-like molecule Consists of two strands four different deoxyribonucleotides DNA Structure 1953 James Watson & Francis Crick ‘The Double Helix’ Basic Structure: "It has not escaped our notice that the specific pairing we have postulated immediately 1. Two polynucleotide strands wound about a suggests a possible copying mechanism for the genetic material.“ common axis (in anti-parallel orientation) Watson, JD and Crick, FHC (1953) Nature 191, 737-738 2. Sugar-phosphate backbone on the exterior / purine and pyrimidine bases on the interior 3. Complementary base-pairing: A with T G with C Naish medical Sciences Fig 2.15b DNA backbone Covalent bonds hold the sugar-phosphate backbone together 3’ to 5’ phospho- diester bonds Directionality in DNA Each DNA strand has distinct directionality and they are in antiparallel orientation 5’ 3’ One end of a DNA strand has a free hydroxyl group or phosphate group at carbon C-5 (5’) of the last 2’-deoxyribose; this is known as the 5’ end The other end has a free hydroxyl or phosphate at C-3 (3’) of the last 2’-deoxyribose; this is the 3’ end Watson-Crick base pairing Major and minor grooves The sugar-phosphate backbones outline two grooves on the surface of the molecule The edges of the DNA bases are exposed to the environment in the grooves Complementarity Specific base-pairing means that the two strands are ‘complementary’ ie. the sequence of bases along one strand ‘reflects’ that of the other strand Genetic information is stored in the sequence of bases By convention, DNA base sequences are usually written from 5’ to 3’ 5’-atccggatcgtcc-3’ 3’-taggcctagcagg-5’ Forces holding DNA together (in order of strength) Covalent – – covalent bonds within nucleotides – phosphodiester bonds along backbone Non-Covalent: – Base stacking van der Waals interaction between the stacked base- pairs is the strongest non-covalent force in the structure – Hydrogen Bonding G – C : 3 hydrogen bonds A – T : 2 hydrogen bonds A region of DNA which is ‘GC-rich’ is more thermodynamically stable Summary Double helix with anti-parallel orientation Sugar-phosphate backbone on exterior Purine/pyrimidine bases on interior Complementary base- pairing A-T (2 H bonds) Introduction to G-C (3-H bonds) genetic analysis 8th Ed. © W.H. Freeman & Co.  Higher Order Structure of Chromosomes Each human cell contains a very large amount of DNA: 3.2 x 109 bp in 23 chromosomes average length of a chromosome : 3.8cm 46 chromosomes in diploid cell (~1.8 m) How do we fit it all in to a cell which is 10 -6 m in diameter? Chromatin structure and organization DNA is complexed with protein so that it can be organised & folded into compact structures for cell-division. The complex is called chromatin. Chromatin consists of about 50% DNA and 50% protein, by weight In fact, uncomplexed, ‘naked’ DNA is abnormal in the cell Heterochromatin – highly compact, no transcription – no mRNA Euchromatin – diffuse, transcription possible Histones The major proteins involved in the organization of chromatin are histones there are 5 types: H1 H2A H2B H3 H4 with a mass of 11 - 21 kDa All have a high content of positively-charged (basic) amino-acids ~25% Lys or Arg, which allows them to interact with the negatively charged DNA The Nucleosome The most basic repeating structural unit of chromatin is the nucleosome Each nucleosome consists of: ~200 bp of DNA (160 - 240 bp) 2 molecules each of H2A, H2B, H3 & H4 1 molecule of H1 is associated The rest of the DNA lies between nucleosomes and is known as ‘linker DNA’ The Nucelosome The histones (except H1) form an octamer core particle ~ 140 bp of DNA winds around the octamer forming ~ 1.75 turns of a left-handed superhelix H1 sits on the outside of the core-particle X-ray Crystal structure of nucleosome Luger K, Mader AW, Richmond RK, Sargent DF, Richmond TJ, "Crystal Structure of the Nucleosome Core Particle at 2.8 Å Resolution", Nature. 1997 Sep 18; 389 (6648): 251-60. Nucleosome Electron micrograph : “beads on a string” Higher levels of chromosome structure The nucleosomes coil to form a helical or ‘solenoid’ structure of 30 nm diameter known as the ‘30 nm fibre’ At this stage, the length of the DNA has been reduced by ~ 40- fold In fully condensed chromosomes, a total 10,000 fold reduction is achieved.... The solenoid loops and these loops are anchored to scaffold proteins, this structure then undergoes further coiling Basic helix → Nucleosome → Chromatin → Scaffold → Loops → Chromosome 1:1 1:6 1:6 1:36 Compaction - 1: 10,000 Classification of chromatin Heterochromatin describes DNA that is supercoiled and condensed; genes within heterochromatin are not transcribed. Euchromatin describes DNA that is readily accessible to transcription factors and can be transcribed into RNA. Methylation of nucleotide bases promotes compaction while acetylation of specific histone residues promotes relaxation. These reversible modifications are important in control of gene expression http://respiratory-research.com/content/figures/1465- Certain regions of DNA are always in the 9921-7-21-1.jpg heterochromatin form (e.g. telomere (ends); centromere; and non-coding regions. Role of chromatin structure in regulating transcription Lysines are positively charged and will interact with negatively charged DNA on neighbouring nucleosomes – causing DNA condensation If acetylated – the positive charge is neutralized Acetylation promotes looser DNA structure and aids transcription Environmental cues activate Histone Acetyl Transferases (HATs) to unwind DNA and allow gene expression When expression of that gene is not required repressors activate- Histone Deacetylases (HDACs) Clinical note Decondensation of Condensed chromatin chromatin - Activation of silenced - Tumour genes suppressor gene - Induction of cellular silencing differentiation/ cell - Cancer death. HDAC inhibitor (Vorinostat) FDA approved for cutaneous T-cell lymphoma (CTCL) Vorinostat has also demonstrated efficacy in the treatment of solid malignancies (colorectal/ breast) in combination with chemotherapy. Human Chromosomes – essential elements Centromeres : Essential for cell division : attachment point for spindle One per chromosome Origins of replication Points along the chromosome where DNA replication/synthesis begins Many per human chromosome Teleomeres Special structures that cap the ends of chromosomes What makes up human DNA? We actually carry 2 genomes : Nuclear and mitochondrial Strachan and Read : Human Molecular Genetics 3 Human genome consists of 21,300* genes -