AP Biology Chapter 16 Notes: I.
Chapter 16: The Molecular Basis of Inheritance a. Overview: i. April 1953 James Watson and Francis Crick great the double helix model of DNA-­‐ deoxyribonucleic acid ii. DNA is the substance of inheritance (RNA is also in some cases) iii. Nucleic acids can direct their own replication from monomers iv. Heredity information is encoded chemically in DNA v. DNA directs the biochemical, anatomical, physiological and to some extent behavior traits. II.
Chapter 16.1: DNA is the Genetic Material: a. The Search for the Genetic Material: Scientific Inquiry i. T.H. Morgan-­‐ shows that genes are located on chromosomes 1. Chromosomes are made of DNA and proteins 2. DNA and proteins are the two candidates for the genetic material b. Evidence That DNA Can Transform Bacteria: (DNA can be traced back to 1923) i. Frederick Griffith-­‐ British Medical Officer ii. Worked with streptococcus pneumonia iii. Griffith’s experiment: he started with two strains of bacteria, a pathogenic strain(encapsulated) and a nonpathogenic strain(non -­‐encapsulated). 1. He injected mice with the two strains in the following manner a. Living pathogenic strain b. Living nonpathogenic strain c. Heat killed pathogenic strain d. Mixture of heat killed pathogenic strain and living non pathogenic strain 2. Results: a. Mice perished b. Mice lived c. Mice lived d. Mice perished 3. Conclusion: the living non-­‐pathogenic strain had been transformed into pathogenic bacteria by some unknown inheritable substance from the dead pathogenic strain 4. Transformation: a change in genotype and phenotype due to the assimilation of external DNA by a cell a. Do not confuse with the transformation of normal cell to a cancerous one 5. 1944: American bacteriologist Oswald Avery and his team announced that DNA was the transforming material. a. Claim was greeted with interest but much skepticism c. Evidence That Viral DNA Can Program Cells: i. Alfred Hershey and Martha Chase used bacteriophages to solve the dilemma of what is considered to be the heritable material: DNA or proteins 1. Bacteriophage T2 infects E. coli bacteria and is widely used in experimentation a. T2 causes the E. coli bacteria to quickly produce and release many copies of T2 ii. Hershey and Chase experiment: 1. Phages were tagged with a radioactive isotope a. Protein portion of virus was tagged with Sulfur-­‐
35 b. DNA portion of virus was tagged with Phosphorus-­‐32 c. The phages were mixed with bacteria 2. A blender was used to agitate the mixture and separate phages outside the bacteria from the bacterial cells 3. Mixture was centrifuged so bacteria formed a pellet at the bottom of test tube 4. The radioactivity was measured in the liquid and the pellet 5. Results: phage proteins remained outside the bacterial cell during infection, while phage DNA entered the cells. a. When cultured the bacterial cells with radioactive DNA released new phages with some radioactive phosphorus 6. Conclusion: DNA, and not protein was T2’s genetic material d. Additional Evidence That DNA is the Genetic Material: i. Erwin Chargaff: biochemist who analyzed the base composition of DNA from a number of different organisms. 1. 1947-­‐ he reports that DNA composition varies from one organisms to another a. 30.3% of human DNA nucleotides have the Base A. b. 26.0% of the bacterial E. coli have Base A c. this diversity has provided more evidence that DNA is the genetic material 2. He also found “peculiar regularity” in the ratios of nucleotide bases within a single species: a. Humans: i. A = 30.3% ii. T = 30.3% iii. C= 19.9% iv. G = 19.5% b. Chargaff’s rules: the equivalences for any given species between the number of A and T bases and the number of G and C bases. 3. Additional evidence: prior to mitosis the eukaryotic cell doubles, then evenly distributes the DNA to the daughter cells a. In a given species, a diploid set of chromosomes has twice as much as a haploid set. e. Building a Structural Model of DNA: i. Working on the three dimensional structure of DNA: 1. Maurice Wilkins and Rosalind Franklin: a. Franklin while working in Wilkins’ lab produced an X-­‐ray diffraction image of DNA b. Franklin also concludes that the sugar-­‐ phosphate backbone was on the outside of the double helix 2. Watson and Crick (1953): a. Watson was able to deduce from Franklin’s image that DNA was helical in shape and then calculate the width of the helix and the nitrogen bases along it, suggesting it was made up of two strands b. Watson and Crick then began building the model to conform to the X-­‐ray measurements 3. The Double Helix: right handed helix approximately 3.4nm in length per turn a. Sugar phosphate backbone: i. Sugar = ribose= 5 carbon sugar ii. Bound to phosphate via phosphodiester linkage iii. Arranged on the edge of the molecule b. Nitrogen bases: hydrophobic so the are arranged towards the center of the molecule away from an aqueous environment i. Purines = double ringed structures 1. Adenine 2. Guanine ii. Pyrimidines = single ringed structures 1. Thymine 2. Cytosine iii. Adenine pairs with Thymine III.
1. Double ring pairs with single ring to maintain width 2. Linked by two hydrogen bonds iv. Cytosine pairs with Guanine 1. Double ring pairs with single ring to maintain width 2. Linked by three hydrogen bonds Chapter 16.2: Many proteins work together in DNA replication and repair. a. Quote from Watson and Crick publication of the DNA model: “It ahs not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material”. b. The Basic Principle: Base Pairing to a Template Strand i. The two strands are complementary, if you cover one side of the DNA molecule, you can still determine its linear sequence by referring to the uncovered strand 1. Each strand stores the information necessary to reconstruct the other c. Models of DNA replication: i. Basic concept: a short segment of DNA has been untwisted into a structure were each strand will serve as a template, new nucleotides are inserted and each daughter strand consists of one parent strand and one new strand ii. Conservative model: two parent strands re-­‐associate after acting as template for new strands, restoring parent double helix iii. Semiconservative model: the two strands of the parental molecule separate and each function’s as a template for synthesis of a new, complementary strand. iv. Dispersive model: Each strand of both daughter molecules contains a mixture of old and newly synthesized DNA d. DNA Replication: A closer look i. E. coli-­‐ single chromosome about 4.6 million base pairs ii. Humans-­‐ 46 chromosomes about 6 billion base pairs 1. Only takes a few hours to replicate DNA 2. One error per 10 billion nucleotides e. Getting started: Origins of Replication i. Origins of replication-­‐ site where DNA replication starts 1. E. coli-­‐ one origin-­‐ DNA replicates in both directions 2. Humans-­‐ hundreds maybe thousands of origins 3. Origin sites are specific a. Unique DNA segments b. Recognized by origin binding proteins which also function in assembling DNA polymerases and other replication enzymes at the sites where replication begins c. Usually involve an A-­‐T rich stretch ii. Replication bubble: unwinding and separation of the DNA strand for DNA to replicate iii. Replication fork: Y-­‐ shaped region at the each end of the replication bubble where new strands of DNA are elongating f. Elongating a New DNA strand: i. DNA polymerase: group of 11 or so enzymes responsible for catalyzing the elongation of the new DNA strand 1. E. coli-­‐ adds nucleotides at the rate of 500/ sec 2. Humans-­‐ adds nucleotides at the rate of 50/sec ii. DNA polymerase I: Removes primer from the 5’ leading strand and replaces it with DNA, adding on to the adjacent 3’ end 1. Also removes the primer from the 5’ end of each fragment and replaces it with DNA, adding on to the 3’ end of the adjacent fragment iii. DNA polymerase III: continuously synthesizes the leading strand, adding onto the primer 1. Adds DNA nucleotides to the 3’ end of the RNA primer 2. Also synthesizes an RNA primer at the 5’ end of each Okazaki fragment iv. Nucleoside triphosphate: (example: ATP-­‐ only difference is in DNA its deoxyribose and ATP its ribose) 1. As the monomer joins the growing end of DNA it loses two phosphate groups g. Antiparallel Elongation: strands are oriented in opposite directions i. DNA polymerase only adds nucleotides in one direction (5’ to 3’) it adds them on the 3’ end 1. Leading strand: DNA polymerase III adds nucleotides continuously in the 5’ to 3’ direction a. In the direction of the replication fork b. Leading strand needs only one primer 2. Lagging strand-­‐ DNA synthesized in a series of segments by DNA Polymerase III away from the replication fork a. Okazaki fragments-­‐ segments of lagging strand i. Each Okazaki fragments needs a primer b. Sugar/phosphate backbone joined together by DNA ligase i. E. coli-­‐ fragments are 1000 – 2000 nucleotides long ii. Humans-­‐ fragments are 100-­‐ 200 nucleotides long iii. DNA ligases close nicks in the phosphodiester
backbone of DNA. Biologically, DNA ligases are iv. essential for the joining of Okazaki fragments
during replication, and for completing shortpatch DNA synthesis occurring in DNA repair
process. There are two classes of DNA
ligases. The first uses NAD+ as a cofactor and
only found in bacteria. The second uses ATP as
a cofactor and found in eukaryotes, viruses and
bacteriophages. The smallest known ATPdependent DNA ligase is the one from the
bacteriophage T7 (at 41KdA). Eukaryotic DNA
ligases may be much larger (human DNA ligase
I is > 100KDA) but they all appear to share some
common sequences and probably structural
motifs. 2+
v. Requires ATP and Mg h. Priming DNA synthesis: i. Primer-­‐ initial nucleotide chain 1. May consist of DNA or RNA a. Initiating the replication of cellular DNA, the primer is a short stretch of RNA with an available 3’ end 2. Catalyzed by primase-­‐ starts the RNA chain from scratch 3. Generally 5 – 10 nucleotides long 4. DNA polymerase III adds nucleotides on the 3’ end 5. Okazaki fragments are primed separately ii. Primase: synthesizes a single RNA primer at the 5’ end of the leading strand 1. Also synthesizes an RNA primer at the 5’ end of each Okazaki fragment i. Other proteins that assist DNA replication: i. Helicase-­‐ untwists the DNA helix at the replication fork ii. Topoisomerase-­‐ helps relieve the strain of tighter twisting from the action helicase 1. Relieves torsional tensions by breaking and rejoining the double strands iii. Single strand binding protein: Binds to and stabilizes single-­‐ stranded DNA until it can be used as a template j. Replicating the Ends of DNA Molecules: i. Small portions of the DNA strand cannot be replicated or repaired by DNA polymerase 1. They can only add nucleotides at the 3’ end 2. Results in the repeated rounds of replication produce shorter and shorter DNA molecules a. Prokaryotes have circular DNA so they do not have this problem ii. Telomeres: nucleotide sequences at the ends of the DNA molecule 1. They do not contain genes 2. Contain a repeated unit a. In humans this repeated unit is TTAGGG b. Number of repetitions varies from 100 to 1000 3. Protects the molecule from the replication process 4. Can trigger cell cycle arrest or apoptosis 5. They postpone the erosion of the genes near the end 6. The shortening of the telomeres may contribute to the aging process iii. Telomerase: catalyzes the lengthening of the telomeres in eukaryotic germ cells restoring their original length