DNA Structure & Replication PDF

Presented by Dr Ghada Khawaja DNA Structure & DNA replication Overview: Life’s Operating Instructions In 1953, James Watson and Francis Crick introduced an elegant double- helical model for the structure of deoxyribonucleic acid, or DNA DNA, the substance of inh...

Presented by Dr Ghada Khawaja DNA Structure & DNA replication Overview: Life’s Operating Instructions In 1953, James Watson and Francis Crick introduced an elegant double- helical model for the structure of deoxyribonucleic acid, or DNA DNA, the substance of inheritance, is the most celebrated molecule of our time Hereditary information is encoded in DNA and reproduced in all cells of the body This DNA program directs the development of biochemical, anatomical, physiological, and (to some extent) behavioral traits Concept 1: DNA is the genetic material, Nucleic acids store and transmit hereditary information The amino acid sequence of a polypeptide is programmed by a unit of inheritance called a gene (or is encoded by a gene) Genes are made of DNA, a nucleic acid The Roles of Nucleic Acids There are two types of nucleic acids: – Deoxyribonucleic acid (DNA) – Ribonucleic acid (RNA) DNA provides directions for its own replication – Thus, as a cell divides, its genetic instructions (DNA) are passed to each daughter cell DNA directs synthesis of messenger RNA (mRNA) and, through mRNA, controls protein synthesis Protein synthesis occurs in ribosomes  DNA -Genes in DNA do not build proteins directly 1 -They work through an intermediary, the Synthesis of second type of nucleic acid, the RNA mRNA in the nucleus mRNA - In the nucleus of a eukaryotic cell, a gene directs the synthesis of an RNA molecule  DNA is transcribed into RNA NUCLEUS - RNA molecules moves out of the nucleus and CYTOPLASM interact with the protein building machinery of the cell called ribosome 2 mRNA - There, the gene’s instructions written in Movement of mRNA into cytoplasm Ribosome nucleic acid language are translated into via nuclear pore protein language, the aa sequence of a polypeptide 3 - P.S: in prokaryotic cells, which lack nuclei, Synthesis of protein both transcription and translation take place within the cytoplasm of the cell Amino Polypeptide acids The Structure of Nucleic Acids Nucleic acids are polymers called polynucleotides Each polynucleotide is made of monomers called nucleotides Each nucleotide consists of a nitrogenous base, a pentose sugar, and a phosphate group The portion of a nucleotide without the phosphate group is called a nucleoside Nucleotide Monomers There are two families of nitrogenous bases: – Pyrimidines (cytosine, thymine, and uracil) have a single six-membered ring – Purines (adenine and guanine) have a six- membered ring fused to a five-membered ring Sugars In DNA, the sugar is deoxyribose; in RNA, the sugar is ribose Deoxyribose (in DNA) Ribose (in RNA) (c) Nucleoside components: sugars 5’ end Nucleotide Polymers Nitrogenous bases Pyrimidines 5 C 3 C Nucleoside Nitrogenous base Cytosine (C) Thymine (T, in DNA) Uracil (U, in RNA) Purines Phosphate group Sugar 5 C (pentose) Adenine (A) Guanine (G) 3 C (b) Nucleotide Sugars 3’ end (a) Polynucleotide, or nucleic acid - Nucleotide polymers are linked together to build a polynucleotide - Adjacent nucleotides are joined by covalent bonds that form between the –OH group on the 3 carbon of one nucleotide and the phosphate Deoxyribose (in DNA) Ribose (in RNA) on the 5 carbon on the next (c) Nucleoside components: sugars - These links create a backbone of sugar-phosphate units with nitrogenous bases as attachments (nitrogenous bases are not part of the backbone) - The sequence of bases along a DNA or mRNA polymer is unique for each gene The DNA Double Helix A DNA molecule has two polynucleotides spiraling around an imaginary axis, forming a double helix In the DNA double helix, the two backbones run in opposite 5 → 3 directions from each other, an arrangement referred to as antiparallel One DNA molecule includes many genes The nitrogenous bases in DNA pair up and form hydrogen bonds: adenine (A) always with thymine (T), and guanine (G) always with cytosine (C) - DNA is a double helix 5' end 3' end - Because of the base pairing rules, the 2 strands of the double helix are said to be complementary, each a Sugar-phosphate predictable counter part of the other backbones (e.g. If a stretch of nucleotides on one strand has the base Base pair (joined by sequence AGCACT, then the same stretch on the other strand must be TCGTGA) hydrogen bonding) -Complementary base pairing is the key to know a cell makes two identical copies of each of its DNA molecules every time Old strands it divides Nucleotide - Thus, the structure of DNA accounts for its function of about to be transmitting genetic information whenever a cell reproduces. added to a new strand - The same base pairing rules (with the 3' end exception that U nucleotides of RNA pair with A nucleotides of DNA) also account for the precise transcription of information from DNA to RNA. 5' end An organism’s genes determine the proteins and thus the structures and functions of its New body. strands 3' end 5' end 5' end 3' end Concept 2: Many proteins work together in DNA replication and repair The relationship between structure and function is manifest in the double helix Watson and Crick noted that the specific base pairing suggested a possible copying mechanism for genetic material The Basic Principle: Base Pairing to a Template Strand Since the two strands of DNA are complementary, each strand acts as a template for building a new strand in replication In DNA replication, the parent molecule unwinds, and two new daughter strands are built based on base-pairing rules DNA Replication: A Closer Look The copying of DNA is remarkable in its speed and accuracy More than a dozen enzymes and other proteins participate in DNA replication Getting Started Replication begins at particular sites called origins of replication, where the two DNA strands are separated, opening up a replication “bubble” A eukaryotic chromosome may have hundreds or even thousands of origins of replication Replication proceeds in both directions from each origin, until the entire molecule is copied (a) Origin of replication in an E. coli cell (b) Origins of replication in a eukaryotic cell Origin of Double-stranded Parental (template) strand Origin of replication DNA molecule replication Daughter (new) strand Parental (template) Daughter (new) strand strand Replication Double- fork stranded DNA molecule Replication bubble Bubble Replication fork Two daughter DNA molecules Two daughter DNA molecules 0.25 m 0.5 m  At the end of each replication bubble is a replication fork, a Y-shaped region where new DNA strands are elongating Helicases are enzymes that untwist the double helix at the replication forks Single-strand binding proteins bind to and stabilize single-stranded DNA Topoisomerase corrects “overwinding = overtwisting” ahead of replication forks by breaking, swiveling, and rejoining DNA strands Primase 3 Topoisomerase 5 RNA 3 primer 5 3 Helicase 5 Single-strand binding proteins DNA polymerases cannot initiate synthesis of a polynucleotide; they can only add nucleotides to the 3 end The initial nucleotide strand is a short RNA primer An enzyme called primase can start an RNA chain from scratch and adds RNA nucleotides one at a time using the parental DNA as a template The primer is short (5–10 nucleotides long), and the 3 end serves as the starting point for the new DNA strand Synthesizing a New DNA Strand Enzymes called DNA polymerases catalyze the elongation of new DNA at a replication fork Most DNA polymerases require a primer and a DNA template strand The rate of elongation is about 500 nucleotides per second in bacteria and 50 per second in human cells Each nucleotide that is added to a growing DNA strand is a nucleoside triphosphate dATP supplies adenine to DNA and is similar to the ATP of energy metabolism The difference is in their sugars: dATP has deoxyribose while ATP has ribose As each monomer of dATP joins the DNA strand, it loses two phosphate groups as a molecule of pyrophosphate Antiparallel Elongation The antiparallel structure of the double helix affects replication DNA polymerases add nucleotides only to the free 3end of a growing strand; therefore, a new DNA strand can elongate only in the 5to 3direction Along one template strand of DNA, the DNA polymerase synthesizes a leading strand continuously, moving toward the replication fork Overview Leading Origin of replication Lagging strand strand Primer Lagging Leading strand strand Origin of Overall directions replication of replication 3 5 5 RNA primer 3 3 Sliding clamp DNA pol III Parental DNA 5 3 5 5 3 3 5  Overview Leading strand Origin of replication Lagging strand Primer Lagging Leading strand strand Overall directions of replication Origin of replication 3 5 5 RNA primer 3 3 Sliding clamp DNA pol III Parental DNA 5 3 5 5 3 3 5  3 Overview 5 3 Leading Origin of replication Lagging Template strand strand strand RNA primer 5 for fragment 1 Lagging strand 3 2 1 5 Leading 1 3 strand Overall directions 5 of replication 3 Okazaki fragment 1 5 1 RNA primer 3 for fragment 2 5 5 Okazaki 3 fragment 2 2 1 3 5 5 3 2 1 3 5 5 3 2 1 3 5 Overall direction of replication Overview Leading Origin of replication Lagging strand strand Lagging strand 2 1 Leading strand Overall directions of replication  3 5 3 Template strand 5 To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase 3 5 3 Template strand 5 3 RNA primer for fragment 1 5 1 3 5 To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase  3 5 3 Template strand 5 3 RNA primer for fragment 1 5 1 3 5 3 Okazaki fragment 1 5 1 3 5 To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase 3 5 3 Template strand 5 3 RNA primer for fragment 1 5 1 3 5 3 Okazaki fragment 1 5 1 RNA primer 3 for fragment 2 5 5 3 2 Okazaki fragment 2 1 3 5 To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase  3 5 3 Template strand 5 3 RNA primer for fragment 1 5 1 3 5 3 Okazaki fragment 1 5 1 RNA primer 3 for fragment 2 5 5 3 2 Okazaki fragment 2 1 3 5 5 3 2 1 3 5 5 3 3 5 3 Template strand 5 3 RNA primer for fragment 1 5 1 3 5 3 Okazaki fragment 1 5 1 RNA primer 3 for fragment 2 5 5 3 2 Okazaki fragment 2 1 3 5 5 3 2 1 3 5 5 3 2 1 3 5 Overall direction of replication  DNA pol III synthesizes leading strand continuously 3 5 Parental DNA DNA pol III starts DNA synthesis at 3 end of primer, Origin of 5 continues in 5 3 direction replication 3 5 Lagging strand synthesized in short Okazaki fragments, Helicase later joined by DNA ligase Primase synthesizes 3 a short RNA primer 5 DNA pol I replaces the RNA primer with DNA nucleotides The DNA Replication Complex The proteins that participate in DNA replication form a large complex, a “DNA replication machine” The DNA replication machine may be stationary during the replication process  DNA pol III Parental DNA Leading strand 5 5 3 3 3 3 5 5 Connecting Helicase protein 3 5 Lagging DNA strand 3 Lagging strand template pol III 5 Proofreading and Repairing DNA DNA polymerases proofread newly made DNA, replacing any incorrect nucleotides In mismatch repair of DNA, repair enzymes correct errors in base pairing DNA can be damaged by exposure to harmful chemical or physical agents such as cigarette smoke and X-rays; it can also undergo spontaneous changes In nucleotide excision repair, a nuclease cuts out and replaces damaged stretches of DNA  5 3 3 5 Nuclease 5 3 3 5 DNA polymerase 5 3 3 5 DNA ligase 5 3 3 5 Evolutionary Significance of Altered DNA Nucleotides Error rate after proofreading repair is low but not zero Sequence changes may become permanent and can be passed on to the next generation These changes (mutations) are the source of the genetic variation upon which natural selection operates Replicating the Ends of DNA Molecules Limitations of DNA polymerase create problems for the linear DNA of eukaryotic chromosomes Because DNA polymerases can only add nucleotides to the 3 end of a preexisting polynucleotide, the usual replication machinery provides no way to complete the 5 ends, so repeated rounds of replication produce shorter DNA molecules with uneven ends This is not a problem for prokaryotes, most of which have circular chromosomes 5 Ends of parental Leading strand DNA strands Lagging strand 3 Last fragment Next-to-last fragment Lagging strand RNA primer 5 3 Parental strand Removal of primers and replacement with DNA where a 3 end is available 5 3 Second round of replication 5 New leading strand 3 New lagging strand 5 3 Further rounds of replication Shorter and shorter daughter molecules  Eukaryotic chromosomal DNA molecules have special nucleotide sequences at their ends called telomeres Telomeres do not prevent the shortening of DNA molecules, but they do postpone the erosion of genes near the ends of DNA molecules It has been proposed that the shortening of telomeres is connected to aging If chromosomes of germ cells became shorter in every cell cycle, essential genes would eventually be missing from the gametes they produce An enzyme called telomerase catalyzes the lengthening of telomeres in germ cells The shortening of telomeres might protect cells from cancerous growth by limiting the number of cell divisions There is evidence of telomerase activity in cancer cells, which may allow cancer cells to persist 1 m Concept 3: A chromosome consists of a DNA molecule packed together with proteins The bacterial chromosome is a double-stranded, circular DNA molecule associated with a small amount of protein Eukaryotic chromosomes have linear DNA molecules associated with a large amount of protein In a bacterium, the DNA is “supercoiled” and found in a region of the cell called the nucleoid Chromatin, a complex of DNA and protein, is found in the nucleus of eukaryotic cells Chromosomes fit into the nucleus through an elaborate, multilevel system of packing Nucleosome (10 nm in diameter) DNA double helix (2 nm in diameter) H1 Histone Histones tail Nucleosomes, or “beads on DNA, the double helix Histones a string” (10-nm fiber)