Mr. Anderson: Replication ( AP Biology

Transcription

1 Mr. Anderson: Replication (

2 Copying DNA S Phase Replication occurs during the S-Phase of the cell cycle. Each DNA molecule is used to synthesize identical daughter strands of DNA. The cell must replicate it's DNA so that there will be enough genetic material for new cells formed through mitosis, to each have a correct and identical set of DNA. Replication occurs in the nucleus of Eukaryotic cells and the cytoplasm of Prokaryotic

3 Basic Replication animation Steps involved in Replication: *remember - genetic information is stored in the order of the bases in the rungs of DNA. So in order to replicate the information the double stranded DNA molecule must open so the order of the bases can be replicated.

4 Site where it starts = ORIGIN of REPLICATION - DNA strands are separated, opening up a replication 'bubble'. Place where nucleotides add = REPLICATION FORK. Note that there is a replication fork at each end of the bubble. Prokaryote- single starting spot Eukaryotes-multiple sites Link - Bacterial replication

5 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 Recent studies support a model in which DNA polymerase molecules reel in parental DNA and extrude newly made daughter DNA molecules 2011 Pearson Education, Inc.

6 Figure DNA pol III Parental DNA Leading strand Connecting protein Helicase DNA pol III Lagging strand Lagging strand template

7 The DNA Replication Complex includes: 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 2011 Pearson Education, Inc.

8 DNA POLYMERASE III reads code strand in 3 5 direction builds a new strand in 5 3 direction adds on to 3 end of sugar in previous nucleotide

9 Energy of Replication Splitting phosphates from nucleotide triphosphate subunits provides energy for the replications reaction The nucleotides arrive as nucleoside triphosphates DNA bases with P P P P-P-P = energy for bonding DNA bases arrive with their own energy source for bonding bonded by enzyme: DNA polymerase III ATP GTP TTP CTP

10 Energy of Replication Where does energy for bonding usually come from? You remember ATP! Are there other energy ways nucleotides? to get energy You out of bet! it? We come with our own energy! energy energy GTP TTP CTP ATP modified nucleotide And we leave behind a nucleotide! ADP AMP GMP TMP CMP

11 Adding Complementary Bases DNA Polymerase III Where s the ENERGY for the bonding come from?

12 DNA POLYMERASE CAN T START A CHAIN by itself; can only add nucleotides to end of an existing DNA/RNA Chain, therefore, a new DNA strand can elongate only grows energy DNA Polymerase III energy DNA Polymerase III energy DNA Polymerase III DNA Polymerase III energy

13 Replication Enzymes HELICASE- untwists double helix to open strands at replication forks TOPOISOMERASE- relieves strain caused by untwisting, breaking and rejoining DNA strands. SINGLE-STRAND BINDING PROTEINSstabilize unpaired strands to hold them open Link: DNA REPLICATION FORK helicase single-stranded binding proteins replication fork

14 Replication Enzymes PRIMASE-starts segment by adding a primer made up of 5-10 RNA nucleotides, and the 3' end serves as a starting point for the new DNA strand. *Evolutionary significance - Since a primer is made from RNA and can be used as a template for DNA, it suggests that RNA existed before DNA.

15 Replication Enzymes DNA POLYMERASE I removes RNA primers and replaces them with DNA bases by adding to the 3 end of the previous fragment LIGASE-joins Okazaki fragments together to make a continuous copied strand

16 need primer bases to add on to no energy to bond energy energy energy energy energy ligase energy energy

17 Okazaki Leading & Lagging strands Limits of DNA polymerase III can only build onto end of an existing DNA strand ligase Lagging strand growing replication fork Lagging strand Okazaki fragments joined by ligase spot welder enzyme DNA polymerase III Leading strand Leading strand continuous synthesis

18 LEADING STRAND (Original DNA strand runs 3 5 ) copies toward the replication fork PRIMASE adds RNA primer to start chain DNA POLYMERASE III adds nucleotides in 5 3 direction (referring to new molecule)

19 Replication fork / Replication bubble DNA polymerase III leading strand lagging strand growing replication fork lagging strand leading strand leading strand lagging strand growing replication fork

20 LAGGING STRAND (Original DNA strand runs 5 3 ) copies away from replication fork PRIMASE adds RNA primers at various spots as fork opens DNA POLYMERASE III adds nucleotides in 5 3 (referring to new molecule) direction in short segments= OKAZAKI FRAGMENTS

21 Starting DNA synthesis: RNA primers Limits of DNA polymerase III can only build onto end of an existing DNA strand growing replication fork DNA polymerase III primase RNA RNA primer built by primase serves as starter sequence for DNA polymerase III

22 Replacing RNA primers with DNA DNA polymerase I removes sections of RNA primer and replaces with DNA nucleotides DNA polymerase I growing replication fork ligase RNA But DNA polymerase I still can only build onto end of an existing DNA strand Link: HOW NUCLEOTIDES ARE ADDED

23 Chromosome erosion All DNA polymerases can only add to end of an existing DNA strand Houston, we have a problem! DNA polymerase I growing replication fork DNA polymerase III RNA Loss of bases at ends in every replication chromosomes get shorter with each replication limit to number of cell divisions?

24 Primer removed but can t be replaced with DNA because no 3 end available for DNA POLYMERASE TELOMERES & TELOMERASE IMPORTANT: Because DNA polymerase can t fill in last section when primer is removed from lagging strand, the code shortens with each replication (usually only a problem for Eukaryotes since prokaryotes have circular chromosomes) Image from: AP BIOLOGY by Campbell and Reese 7 th edition

25 Eukaryotic chromosomal DNA molecules have special nucleotide sequences at their ends called telomeres. TELOMERE sequences at ends of chromosomes to help postpone the erosion of essential information in code / genes with each replication. it has been proposed that the shortening of telomeres may play a role in a aging and cancer Link 1: TELOMERASE = enzyme that lengthens telomeres found in eukaryotic germ cells that divide frequently to produce gametes Link 2: TELOMEREShttp://stemcells.nih.gov/info/scireport/appendixC.asp Link 3: ANIMATION

26 PROOFREADING & REPAIR Mistakes in final DNA: 1 in 10 billion Mistakes in initial base pairing during replication 1 in 100,000 DNA POLYMERASE proofreads each base as it s added & fixes errors Errors can come from proofreading mistakes that are not caught OR environmental damage (Ex: X-rays, UV light, chemical mutagens/ carcinogens)

27 NUCLEOTIDE EXCISION REPAIR Cells continually monitor DNA and make repairs 1. NUCLEASES- DNA cutting enzymes remove errors 2. DNA POLYMERASE fills in gap using complimentary strand 3. LIGASE seals ends

28 Ex: THYMINE DIMERS = joins THYMINES in same strand damage caused by UV light can be repaired

29 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

30 Replication fork DNA polymerase I DNA polymerase III Okazaki fragments lagging strand 5 ligase 3 5 primase 3 SSB 3 helicase leading strand direction of replication DNA polymerase III SSB = single-stranded binding proteins

31 DNA polymerases DNA polymerase III 1000 bases/second! main DNA builder DNA polymerase I 20 bases/second editing, repair & primer removal DNA polymerase III enzyme Thomas Kornberg Arthur Kornberg 1959

32 Replication Summary. Go trough the following animation to review the entire process of replication: Link: ns/replication1.swf Be sure to complete the four parts a) Replication fork b) Fork with proteins c) Concerted replication d) Trombone model