Mead AP Biology
Lap 5 Molecular Genetics
Chapters 16-17
16.1 DNA as Genetic Material
A. Search for genetic material
1. Evidence that DNA can transform bacteria
 Griffith 1928, Fig 16.2
 Streptococcus pnemonia
 Transformation
 Avery, McCarty, Macleod 1944
 Identify DNA as transformation agent, NOT proteins
2. Evidence that viral DNA can program cells
 Bacteriophage, Fig 16.3
 Hershey and Chase, 1952
 Fig 16.4 Experiment
 T2 phages
 Radioactive tags
3. Additional Evidence
 Chargaff 1947
 Knew DNA components
 Base composition
 Varies by species
 Constant ratios
 A=T
 C=G
B. Building Structural Model of DNA
1. Relate structure to function
 Early 1950s, knew basic structure, Fig 16.5
 Still looking for 3D structure
2. Wilkins and Franklin, Pauling
 X-ray diffraction
 Helical, 2 or 3 strands, Fig 16.6
3. Watson and Crick Build 3D model
 Double helix
 10 base pairs per turn
 Hydrophobic bases on inside
 Base pairing according to Chargaff’s rule
16.2 DNA Replication and Repair
A. Basic Principle
◦ Steps, Fig 16.9
◦ 3 Models, Fig 16.10
1. Conservative
2. Semi-conservative
3. Dispersive
◦ Meselson-Stahl prove Semi-conservative
 Fig 16.11
 Original DNA: N15
 New DNA: N14
 Centrifuge after 1 and 2 replications
B. DNA Replication Details
1. Origins of replication, Fig 16.12
 Def
 Initiation proteins recognize and bind
 Replication “bubble”
 Replication fork
2. Elongating new strand
 DNA polymerase
 Energy provided by 2 phosphate groups
 Fig 16. 13
3. Antiparallel Elongation
◦ Strands run antiparallel, Fig 16.13
 5’ end: phosphate on top
◦ DNA polymerase III adds to 3’ only
◦ Leading strand, Fig 16.14
◦ Lagging strand
 Okazaki fragments
 Ligase
4. Priming DNA synthesis, Fig 16.15
◦ Need primer
 Def
◦ Primase
◦ Leading strand:
 Need 1
◦ On lagging strand:
 Need for each Okazaki fragment
 Replace with DNA by DNA polymerase I
 Ligase
5. Other proteins assist in replication, Fig 16.16
◦ Helicase
◦ Single strand binding protein
◦ Topoisomerase
6. Summary, Table 16.1 and Fig 16.16
C. Proofreading and Repairing DNA
◦ Mismatch repair
◦ Repair damage caused by chemical/physical agents
◦ Nucleotide Excision Repair
 Fig 16.17
 Nuclease
 Polymerase
 Ligase
D. Replicating ends of DNA
◦ DNA polymerase can only add to 3’
 Prokaryote: circular DNA no effect
 Eukaryote: can’t replace primer
◦ Telomeres
 Def
 TTAGGG
◦
Telomerase
 Enzyme function
 Germ cells
 Link to aging
 Cancer
17.1 Genes Specify Proteins
A. Evidence from the study of metabolic disorders
◦ 1909 Garrod: Genes dictate phenotypes through enzymes
 Alkaptonuria
◦ 1930s: Fruit fly eye color
◦ Beadle and Tatum: Work with bread mold to create one gene-one enzyme hypothesis
 Metabolic pathway needed to make amino acids
 Precursor Ornithine Citrulline Arginine
A
B
C
 Each reaction catalyzed by an enzyme
 If mutation at A: must add Orn
 If mutation at B: must add Cit
B. Basic principles of Transcription and Translation
 Fig 17.3
1. Genes (DNA) provide instructions for proteins
2. RNA connects DNA to protein
 RNA structure
3. Transcription: synthesis of mRNA
4. Translation: synthesis of polypeptide chain
 RNA base sequence to amino acid sequence
 tRNA
 rRNA
5. Eukaryotic cells add RNA processing: make pre mRNA
C. The Genetic Code
◦ Codons: triplets of bases
◦ DNA template mRNA codon amino acids: Fig 17.4
◦ Cracking the code, Fig 17.5
 Deciphered by mid 1960s
 Start/methionine: AUG
 Stop: UAA / UAG / UGA
 Multiple versions of each amino acid keep error low
 “Reading frame”
D. DNA code evolution
◦ Nearly universal
◦ Transplant genes between species
◦ Fig 17.6: firefly gene inserted into tobacco
◦ Uses
17.2 Transcription: Details
A. RNA polymerase
◦ Function
◦ Transcription unit
◦ Bacteria vs Eukaryote
◦ Promoter
 In Euk: TATA box
B. 3 Stages of Transcription, Fig 17.7
 Initiation, Fig 17.8
 Transcription factors recognize TATA promoter
 RNA polymerase
 Transcription initiation complex
 Elongation, Fig 17.7
 Termination
 Prokaryotes
 Terminator sequence
 End of transcription unit
 End of gene
 Release mRNA and RNA polymerase
 Eukaryotes
 Polyadenylation sequence
 AAUAAA
 RNA polymerase releases later
 Move on to RNA processing
17.3 Eukaryotic cells modify RNA
A. Alteration of mRNA ends, Fig 17. 9
◦ 5’ cap: Modified Guanine
◦ Poly-A tail
◦ Functions
B. Split genes and RNA splicing
◦ RNA splicing
◦ Exons and Introns
◦ Introns removed
◦ Spliceosomes, Fig 17.11
 snRNP and proteins
◦ Ribozymes
◦ Importance of Introns
17.4 Translation: Details
A. Translation: Overview
◦ Fig 17.13
B. Molecules involved in translation
◦ Transfer RNA, Fig 17.14
 tRNA
 Anticodon
 Unique for each amino acid
 Structure
 tRNA “wobble”
◦ Aminoacyl-tRNA synthetase, Fig 17.15
 Uses ATP for energy
◦ Ribosomes, Fig 17.16
 2 subunits
 Structure: Binding sites
 1 for mRNA and 3 for tRNA: E,P, A
 Function
C. 3 Steps of Building a Polypeptide
1. Initiation, Fig 17.17
 Create Translation Initiation Complex
 Small ribosomoal subunit
 mRNA
 Initiator tRNA (methionine): into P site
 Large ribosomal subunit
 Initiation factors (proteins) bring it all together
 Requires GTP for energy
2. Elongation, Fig 17.18
 Codon recognition: A site
 Peptide bond formation
 Translocation
 Requires elongation factors
 Requires energy: GTP
3. Termination, Fig 17.19
 Stop codon reaches P site
 Release factor enters A site
 Breaks bond of tRNA in P site and last amino acid
 Components disassemble
 Polyribosomes, Fig 17.20
 Simultaneous translation
D. Completing and Targeting Protein
◦ Folding and Modifications
◦ Targeting to specific locations
 Fig 17.21
 Start translation in a free cytoplasmic ribosome
 SRP halts translation
 Moves to ER
17.5 RNA Roles in Cell: Table 17.1
17.6 Point Mutations Affect Protein
A. Point mutation
◦ Def
◦ Types
 Substitutions, Fig 17.24
 Silent
 Missense
 Nonsense
 Example: Sickle Cell Anemia, Fig 17.23
 Insertion/Deletion
 Fig 17.25
 Frame-shift mutations
B. Mutagens
◦ Spontaneous mutations
◦ Mutagens def
◦ Ames test
Summary of transcription and translation, Fig 17.26