AP Biology DNA The Genetic Material

AP Biology Scientific History  The march to understanding that DNA is the genetic material  T.H. Morgan (1908)  Frederick Griffith (1928)  Avery, McCarty & MacLeod (1944)  Erwin Chargaff (1947)  Hershey & Chase (1952)  Watson & Crick (1953)  Meselson & Stahl (1958)

AP Biology Chromosomes related to phenotype  T.H. Morgan  working with Drosophila  fruit flies  associated phenotype with specific chromosome  white-eyed male had specific X chromosome 1908 | 1933

AP Biology Genes are on chromosomes  Morgan’s conclusions  ________________________  but is it the protein or the DNA of the chromosomes that are the genes?  initially proteins were thought to be genetic material… Why? 1908 | 1933 What ’ s so impressive about proteins?!

AP Biology The “Transforming Principle” 1928  Frederick Griffith  Streptococcus pneumonia bacteria  was working to find cure for pneumonia  harmless live bacteria (“rough”) mixed with heat-killed pathogenic bacteria (“smooth”) causes fatal disease in mice  a substance passed from dead bacteria to live bacteria to change their phenotype  _____________________________

AP Biology The “Transforming Principle” Transformation = change in phenotype something in heat-killed bacteria could still transmit disease-causing properties live pathogenic strain of bacteria live non-pathogenic strain of bacteria mice diemice live heat-killed pathogenic bacteria mix heat-killed pathogenic & non-pathogenic bacteria mice livemice die A.B. C. D.

AP Biology DNA is the “Transforming Principle”  Avery, McCarty & MacLeod  purified both DNA & proteins separately from Streptococcus pneumonia bacteria  which will transform non-pathogenic bacteria?  injected protein into bacteria  no effect  injected DNA into bacteria  transformed harmless bacteria into virulent bacteria 1944 What ’ s the conclusion? mice die

AP Biology Oswald AveryMaclyn McCartyColin MacLeod Avery, McCarty & MacLeod 1944 | ??!!  Conclusion  ___________________________________________ ___________________________________________

AP Biology Confirmation of DNA  Hershey & Chase  classic “blender” experiment  worked with bacteriophage  viruses that infect bacteria  grew phage viruses in 2 media, radioactively labeled with either  ____________________  infected bacteria with labeled phages 1952 | 1969 Hershey Why use S ulfur vs. P hosphorus?

AP Biology Protein coat labeled with 35 S DNA labeled with 32 P bacteriophages infect bacterial cells T2 bacteriophages are labeled with radioactive isotopes S vs. P bacterial cells are agitated to remove viral protein coats 35 S radioactivity found in the medium 32 P radioactivity found in the bacterial cells Which radioactive marker is found inside the cell? Which molecule carries viral genetic info? Hershey & Chase

AP Biology

Blender experiment  Radioactive phage & bacteria in blender  ___________________  radioactive proteins stayed in supernatant  therefore viral protein did NOT enter bacteria  ___________________  radioactive DNA stayed in pellet  therefore viral DNA did enter bacteria  ___________________________________ Taaa-Daaa!

AP Biology Hershey & Chase Alfred HersheyMartha Chase 1952 | 1969 Hershey

AP Biology Chargaff  DNA composition: “________________”  varies from species to species  all 4 bases not in equal quantity  bases present in characteristic ratio  humans: A = 30.9% T = 29.4% G = 19.9% C = 19.8% 1947 That ’ s interesting! What do you notice? Rules A = T C = G

AP Biology Structure of DNA  Watson & Crick  ___________________________________  other leading scientists working on question:  Rosalind Franklin  Maurice Wilkins  Linus Pauling 1953 | 1962 Franklin WilkinsPauling

AP Biology Watson and Crick 1953 article in Nature CrickWatson

AP Biology Rosalind Franklin ( )

AP Biology But how is DNA copied?  Replication of DNA  base pairing suggests that it will allow each side to serve as a template for a new strand “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.”— Watson & Crick

AP Biology Models of DNA Replication  Alternative models  become experimental predictions semiconservative Can you design a nifty experiment to verify? 1 2 P semiconservative

AP Biology Semiconservative replication  Meselson & Stahl  label “parent” nucleotides in DNA strands with heavy nitrogen = 15 N  label new nucleotides with lighter isotope = 14 N “The Most Beautiful Experiment in Biology” 1958 parentreplication Make predictions … 15 N parent strands 15 N/ 15 N

AP Biology Predictions 1st round of replication conservative ________ semi- conservative dispersive conservative semi- conservative dispersive 2nd round of replication 15 N parent strands 15 N/ 15 N 1 2 P ________

AP Biology Franklin Stahl Matthew Meselson Franklin Stahl Meselson & Stahl

AP Biology Scientific History  March to understanding that DNA is the genetic material  T.H. Morgan (1908)  ___________________________________________________  Frederick Griffith (1928)  ___________________________________________________  Avery, McCarty & MacLeod (1944)  ___________________________________________________  Erwin Chargaff (1947)  ___________________________________________________  Hershey & Chase (1952)  ___________________________________________________  Watson & Crick (1953)  ___________________________________________________  Meselson & Stahl (1958)  ___________________________________________________

AP Biology proteinRNA The “Central Dogma” DNA translation replication  Flow of genetic information in a cell translation

AP Biology Science …. Fun Party Time! Any Questions??

AP Biology DNA Replication

AP Biology Watson and Crick 1953 article in Nature

AP Biology Double helix structure of DNA “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.”Watson & Crick

AP Biology Directionality of DNA  You need to number the carbons!  it matters! OH CH 2 O PO 4 N base ribose nucleotide This will be IMPORTANT!!

AP Biology The DNA backbone  Putting the DNA backbone together  refer to the 3 and 5 ends of the DNA  the last trailing carbon OH O PO 4 base CH 2 O base O P O C O –O–O CH Sounds trivial, but … this will be IMPORTANT!!

AP Biology Anti-parallel strands  Nucleotides in DNA backbone are bonded from phosphate to sugar between 3 & 5 carbons  DNA molecule has “direction”  complementary strand runs in opposite direction

AP Biology Bonding in DNA ….strong or weak bonds? How do the bonds fit the mechanism for copying DNA? covalent phosphodiester bonds hydrogen bonds

AP Biology Base pairing in DNA  ________________  ______________  ________________  ______________  Pairing  ______________  2 bonds  ______________  3 bonds

AP Biology Copying DNA  Replication of DNA  base pairing allows each strand to serve as a template for a new strand  new strand is 1/2 parent template & 1/2 new DNA  ____________________ copy process

AP Biology DNA Replication  Large team of enzymes coordinates replication Let ’ s meet the team …

AP Biology Replication: 1st step  Unwind DNA  __________________  unwinds part of DNA helix  stabilized by _____________________________ replication fork helicase I ’ d love to be helicase & unzip your genes … single-stranded binding proteins

AP Biology DNA Polymerase III Replication: 2nd step But … We ’ re missing something! What? Where ’ s the ENERGY for the bonding!  Build daughter DNA strand  add new complementary bases  ___________________

AP Biology energy ATP GTPTTPATP Energy of Replication Where does energy for bonding usually come from? ADPAMPGMPTMPAMP modified nucleotide We come with our own energy! And we leave behind a nucleotide! You remember ATP! Are there other ways to get energy out of it?

AP Biology Energy of Replication  The nucleotides arrive as nucleosides  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: ________________________ ATPGTPTTPCTP

AP Biology  Adding bases  can only add nucleotides to 3 end of the growing DNA strand  need a primer nucleotide to bond to  _________________ DNA Polymerase III Replication energy B.Y.O. ENERGY! The energy rules the process 5

AP Biology no energy to bond 

AP Biology energy ligase

AP Biology Limits of DNA polymerase III  can only build onto 3 end of an existing DNA strand Leading & Lagging strands Leading strand Lagging strand Okazaki fragments ligase Okazaki __________________  continuous synthesis ___________________  __________________  “spot welder” enzyme DNA polymerase III  3 5 growing replication fork

AP Biology DNA polymerase III Replication fork / Replication bubble leading strand lagging strand leading strand lagging strand leading strand growing replication fork growing replication fork lagging strand 5 3

AP Biology DNA polymerase III ______________________  built by ________________  serves as starter sequence for DNA polymerase III Limits of DNA polymerase III  can only build onto 3 end of an existing DNA strand Starting DNA synthesis: RNA primers growing replication fork primase RNA

AP Biology ______________________  removes sections of RNA primer and replaces with DNA nucleotides But DNA polymerase I still can only build onto 3 end of an existing DNA strand Replacing RNA primers with DNA growing replication fork DNA polymerase I RNA ligase

AP Biology Loss of bases at 5 ends in every replication  chromosomes get shorter with each replication  limit to number of cell divisions? DNA polymerase III DNA polymerases can only add to 3 end of an existing DNA strand Chromosome erosion growing replication fork DNA polymerase I Houston, we have a problem!

AP Biology Repeating, non-coding sequences at the end of chromosomes = protective cap  limit to ~50 cell divisions ____________________  enzyme extends telomeres  can add DNA bases at 5 end  different level of activity in different cells  high in stem cells & cancers -- Why? telomerase Telomeres growing replication fork TTAAGGG

AP Biology Replication fork 3’ 5’ 3’ 5’ 3’ 5’ direction of replication

AP Biology 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 Arthur Kornberg 1959 Roger Kornberg 2006

AP Biology Editing & proofreading DNA  1000 bases/second = lots of typos!  DNA polymerase I  proofreads & corrects typos  repairs mismatched bases  removes abnormal bases  repairs damage throughout life  reduces error rate from 1 in 10,000 to 1 in 100 million bases

AP Biology Fast & accurate!  It takes E. coli <1 hour to copy 5 million base pairs in its single chromosome  divide to form 2 identical daughter cells  Human cell copies its 6 billion bases & divide into daughter cells in only few hours  remarkably accurate  only ~1 error per 100 million bases  ~30 errors per cell cycle

AP Biology What does it really look like?

AP Biology Any Questions??