1. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Review When the hydrogen bonds are broken between amino acids in a protein: 1)What structure level are hydrogen bonds located? 2)When the hydrogen bonds are broken, the protein is called? 3)Do enzymes work when hydrogen bonds broken?

2. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings PowerPoint Lectures for Biology, Seventh Edition Neil Campbell and Jane Reece Lectures by Chris Romero Chapter 16 The Molecular Basis of Inheritance

3. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings History of DNA In 1953, James Watson and Francis Crick Described the DOUBLIX HELIX STRUCTURE of DNA Figure 16.1

4. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings What is DNA? DNA, the substance of inheritance DNA directs the development of many different traits by using a chemical language Terms 1.Chromosome – one molecule of DNA & associated protein 2.Genome – collection of ALL of the DNA in a given cell (Humans have 46 chromosomes in genome) 3.GENE – portion of DNA that encodes a single protein

5. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Evidence that DNA is genetic material Scientists first worked out by studying bacteria and the viruses that infect them 1.Griffith studied Streptococcus pneumoniae (bacteria that causes pneumonia in mammals) – He used a pathogenic strain (called S b/c it’s encapsulated) and a nonpathogenic strain (called R b/c it’s not protected.

6. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Griffith found that when he mixed heat-killed remains of the pathogenic strain – With living cells of the nonpathogenic strain, some of these living cells became pathogenic – He called this phenomenon TRANSFORMATION Bacteria of the “S” (smooth) strain of Streptococcus pneumoniae are pathogenic because they have a capsule that protects them from an animal’s defense system. Bacteria of the “R” (rough) strain lack a capsule and are nonpathogenic. Frederick Griffith injected mice with the two strains as shown below: Griffith concluded that the living R bacteria had been transformed into pathogenic S bacteria by an unknown, heritable substance from the dead S cells. EXPERIMENT RESULTS CONCLUSION Living S (control) cells Living R (control) cells Heat-killed (control) S cells Mixture of heat-killed S cells and living R cells Mouse diesMouse healthy Mouse dies Living S cells are found in blood sample. Figure 16.2

7. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Evidence that DNA is genetic material 2. Viruses that infect bacteria, bacteriophages – Are widely used as tools by researchers in molecular genetics Figure 16.3 Phage head Tail Tail fiber DNA Bacterial cell 100 nm

8. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Evidence that DNA is genetic material 3. Chargaff analyzed the base composition of DNA Concluded that DNA composition varies from one species to the next (evidence of diversity) Chargaff’s ratio rule’s for HUMANS: A: 30.3%G: 19.5% T: 30.3% C: 19.9%

9. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Evidence of the Structure of DNA 1.Wilkins & Franklin used a technique called X-ray crystallography to study the structure (a) Rosalind Franklin Franklin’s X-ray diffraction Photograph of DNA (b) Figure 16.6 a, b

10. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Franklin had concluded that DNA – Composed of two antiparallel sugar-phosphate backbones, with the nitrogenous bases paired in the molecule’s interior The nitrogenous bases – Are paired in specific combinations: adenine with thymine, and cytosine with guanine

11. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings O –O–O O OH O –O–O O O H2CH2C O –O–O O O H2CH2C O –O–O O O O O O T A C G C A T O O O CH 2 O O–O– O O 5 end Hydrogen bond 3 end G P P P P O OH O–O– O O O P P O–O– O O O P O–O– O O O P (b) Partial chemical structure H2CH2C 5 end Figure 16.7b O

12. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Structure evidence cont. Figure 16.7a, c C T A A T C G GC A C G A T A T AT T A C T A 0.34 nm 3.4 nm (a) Key features of DNA structure G 1 nm G (c) Space-filling model T 2. Watson and Crick deduced that DNA was a double helix – Through observations of the X-ray crystallographic images of DNA

13. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Watson and Crick reasoned that there must be additional specificity of pairing – Dictated by the structure of the bases Each base pair forms a different number of hydrogen bonds – Adenine and thymine form two bonds, cytosine and guanine form three bonds

14. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings N H O CH 3 N N O N N N NH Sugar Adenine (A) Thymine (T) N N N N Sugar O H N H N H N O H H N Guanine (G) Cytosine (C) Figure 16.8 H

15. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA REPLICATION 1.Definition – manufacture of an EXACT copy of DNA molecule 2.Process of DNA replication A.Parts 1.DNA to be replicated (called parent) 2.Enzymes 1.DNA polymerase (adds nucleotides) 2.Helicase (unwinds the double helix) 3.Single stranded binding proteins (hold DNA open) 4.Primase (begins replication by laying down small “RNA primer”) 5.Ligase (seals the gaps in DNA)

16. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Other Proteins That Assist DNA Replication Helicase, topoisomerase, single-strand binding protein – Are all proteins that assist DNA replication Table 16.1

17. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings In DNA replication – The parent molecule unwinds, and two new daughter strands are built based on base- pairing rules (a) The parent molecule has two complementary strands of DNA. Each base is paired by hydrogen bonding with its specific partner, A with T and G with C. (b) The first step in replication is separation of the two DNA strands. (c) Each parental strand now serves as a template that determines the order of nucleotides along a new, complementary strand. (d) The nucleotides are connected to form the sugar-phosphate backbones of the new strands. Each “daughter” DNA molecule consists of one parental strand and one new strand. A C T A G A C T A G A C T A G A C T A G T G A T C T G A T C A C T A G A C T A G T G A T C T G A T C T G A T C T G A T C Figure 16.9 a–d

18. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA REPLICATION B.Steps 1. Open the complementary DNA strands to expose the nucleotide nitrogenous bases Requires enzyme HELICASE (unwind) & SSB’s (to keep strands apart) 2. Prime the DNA strands Enzyme PRIMASE – forms a very small piece of RNA called RNA primer (DNA polymerase grabs the 3’ OH end of RNA primer)

19. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA REPLICATION 3.Polymerize DNA DNA polymerize aligns appropriate nucleotides across from the template strand (must have RNA primer to use free –OH to start) Rules: 1. NEW strands are built in 5’ to 3’ direction 2. All strands that form pairings must be in opposite direction 3. Leading strand is built continuously & needs only 1 primer 4. Lagging strand is built in segments & needs primer for each individual segment

20. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Overall direction of replication 3 3 3 3 5 3 5 3 5 3 5 3 5 3 5 3 5 3 5 5 1 1 2 1 1 2 5 5 1 2 3 5 Template strand RNA primer Okazaki fragment Figure 16.15 Primase joins RNA nucleotides into a primer. 1 DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. 2 After reaching the next RNA primer (not shown), DNA pol III falls off. 3 After the second fragment is primed. DNA pol III adds DNA nucleotides until it reaches the first primer and falls off. 4 DNA pol 1 replaces the RNA with DNA, adding to the 3 end of fragment 2. 5 DNA ligase forms a bond between the newest DNA and the adjacent DNA of fragment 1. 6 The lagging strand in this region is now complete. 7

21. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA REPLICATION 4.Lagging Strand 1. Helicase opens DNA 2. SSB’s hold them apart 3. Primase lays down RNA primer 4. DNA polymerase forms new DNA strands in segments called OKAZAKI FRAGMENTS (5’ to 3’)

22. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Parental DNA DNA pol Ill elongates DNA strands only in the 5 3 direction. 1 Okazaki fragments DNA pol III Template strand Lagging strand 3 2 Template strand DNA ligase Overall direction of replication One new strand, the leading strand, can elongate continuously 5 3 as the replication fork progresses. 2 The other new strand, the lagging strand must grow in an overall 3 5 direction by addition of short segments, Okazaki fragments, that grow 5 3 (numbered here in the order they were made). 3 DNA ligase joins Okazaki fragments by forming a bond between their free ends. This results in a continuous strand. 4 Figure 16.14 3 5 5 3 3 5 2 1 Leading strand 1 Synthesis of leading and lagging strands during DNA replication

23. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings DNA REPLICATION 5.Fill in the gaps a.Second DNA polymerase “chews out” the RNA primer and fills those areas with DNA b.DNA ligase “seals” the gaps FINAL RESULT 2 double stranded DNA molecules identical to each other & identical to their parental molecule SEMICONSERVATIVE REPLICATION – each daughter is ½ old & ½ new

24. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Figure 16.10 a–c Conservative model. The two parental strands reassociate after acting as templates for new strands, thus restoring the parental double helix. Semiconservative model. The two strands of the parental molecule separate, and each functions as a template for synthesis of a new, comple- mentary strand. Dispersive model. Each strand of both daughter mol- ecules contains a mixture of old and newly synthesized DNA. Parent cell First replication Second replication DNA replication is semiconservative – Each of the two new daughter molecules will have one old strand, derived from the parent molecule, and one newly made strand (a) (b) (c)

25. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Getting Started: Origins of Replication The replication of a DNA molecule – Begins at special sites called origins of replication, where the two strands are separated

26. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings A eukaryotic chromosome – May have hundreds or even thousands of replication origins Replication begins at specific sites where the two parental strands separate and form replication bubbles. The bubbles expand laterally, as DNA replication proceeds in both directions. Eventually, the replication bubbles fuse, and synthesis of the daughter strands is complete. 1 2 3 Origin of replication Bubble Parental (template) strand Daughter (new) strand Replication fork Two daughter DNA molecules In eukaryotes, DNA replication begins at many sites along the giant DNA molecule of each chromosome. In this micrograph, three replication bubbles are visible along the DNA of a cultured Chinese hamster cell (TEM). (b) (a) 0.25 µm Figure 16.12 a, b

27. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Figure 16.16 Overall direction of replication Leading strand Lagging strand Lagging strand Leading strand OVERVIEW Leading strand Replication fork DNA pol III Primase Primer DNA pol III Lagging strand DNA pol I Parental DNA 5 3 4 3 2 Origin of replication DNA ligase 1 5 3 Helicase unwinds the parental double helix. 1 Molecules of single- strand binding protein stabilize the unwound template strands. 2 The leading strand is synthesized continuously in the 5  3 direction by DNA pol III. 3 Primase begins synthesis of RNA primer for fifth Okazaki fragment. 4 DNA pol III is completing synthesis of the fourth fragment, when it reaches the RNA primer on the third fragment, it will dissociate, move to the replication fork, and add DNA nucleotides to the 3 end of the fifth fragment primer. 5 DNA pol I removes the primer from the 5 end of the second fragment, replacing it with DNA nucleotides that it adds one by one to the 3 end of the third fragment. The replacement of the last RNA nucleotide with DNA leaves the sugar- phosphate backbone with a free 3 end. 6 DNA ligase bonds the 3 end of the second fragment to the 5 end of the first fragment. 7 A summary of DNA replication

28. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings 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

29. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Figure 16.17 Nuclease DNA polymerase DNA ligase A thymine dimer distorts the DNA molecule. 1 A nuclease enzyme cuts the damaged DNA strand at two points and the damaged section is removed. 2 Repair synthesis by a DNA polymerase fills in the missing nucleotides. 3 DNA ligase seals the Free end of the new DNA To the old DNA, making the strand complete. 4 In nucleotide excision repair – Enzymes cut out and replace damaged stretches of DNA

30. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Replicating the Ends of DNA Molecules The ends of eukaryotic chromosomal DNA – Get shorter with each round of replication Figure 16.18 End of parental DNA strands Leading strand Lagging strand Last fragmentPrevious fragment RNA primer Lagging strand Removal of primers and replacement with DNA where a 3 end is available Primer removed but cannot be replaced with DNA because no 3 end available for DNA polymerase Second round of replication New leading strand New lagging strand 5 Further rounds of replication Shorter and shorter daughter molecules 5 3 5 3 5 3 5 3 3

31. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings Eukaryotic chromosomal DNA molecules – Have at their ends nucleotide sequences, called telomeres, that postpone the erosion of genes near the ends of DNA molecules Figure 16.19 1 µm

32. Copyright © 2005 Pearson Education, Inc. publishing as Benjamin Cummings If the 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