CH7 key concepts
genetic material: DNA
DNA structure
DNA replication
LECTURE 12: DNA: STRUCTURE & REPLICATION I
CHAPTER 7: KEY CONCEPTS transforming principle in bacteria is DNA experiments with labeled T2 phage establish that
DNA is the hereditary material Watson & Crick DNA model: anti-parallel double
helix = nucleotides linked by phosphodiester bonds; helices are held together by hydrogen bonds between pairs of nitrogenous bases
DNA structure ensures fidelity of replication; base pairing specified by hydrogen bonding
DNA replication is semiconservative; daughter duplex has 1 parental & 1 new strand
DNA replication depends on many enzymes
DNA = GENETIC MATERIAL Griffith (1928), Streptococcus pneumoniae transforming principle in bacteria is DNA
DNA = GENETIC MATERIAL Avery et al. (1944), Streptococcus pneumoniae more proof ... transforming principle is DNA
DNA = GENETIC MATERIAL Hershey & Chase (1952), T2 bacteriophage DNA (not protein) is genetic material
why was the scientific community reluctant to accept DNA (rather than protein) as hereditary material?...
DNA is a simple molecule...
how is complexity of life encoded by such simplicity?
how is information accurately transmitted from one generation to the next?
DNA = GENETIC MATERIAL
DNA STRUCTURE
hydrogen bonded nucleotides on opposite helices
DNA helices are antiparallel
carbons on sugar define ends... 5' and 3'
pyrimidines bond with purines T A C G
DNA STRUCTURE bonding is specific: pyrimidines bond with purines implication: mechanism for accurate replication
DNA STRUCTURE helical structure of DNA
major & minor groves
10Å radius & 20Å diameter
3.4Å between nucleotide base pairs
34Å / 360° turn
10 nucleotide base pairs / 360° turn
DNA STRUCTURE
clues for Watson & Crick (1953) ... X-ray data
& ratios ( A + G ) = ( T + C ) ... Chargaff
DNA REPLICATION model of replication proposed
by Watson & Crick (1953)
parental strand = template
semiconservative model (new double helix has 1 template + 1 new daughter strand)
3 possible models: conservative semiconservative dispersive
DNA REPLICATION plant root cells incubated with [3H] thymidine
colchicine to inhibit spindles (chromosomes do not separate)
autoradiography
DNA REPLICATION explanation of [3H] thymidine incubation / colchicine
treatment / autoradiography at DNA level
DNA REPLICATION harlequin chromosomes of Chinese hamster ovary
cells, incubated with bromodeoxyuridine (BUdR)
chromatidexchange
DNA REPLICATION
semiconservative replication
linear gene arrangement
... chromatid = single DNA molecule + protein matrix
chromatidexchange
DNA REPLICATION model of replication proposed
by Watson & Crick (1953)
parental strand = template
semiconservative model (new double helix has 1 template + 1 new daughter strand)
DNA REPLICATION model of replication proposed
by Watson & Crick (1953)
parental strand = template
semiconservative model (new double helix has 1 template + 1 new daughter strand)
replication fork
DNA REPLICATION
replication fork ? bacteria incubated with [3H] thymidine... daughter molecules 1 “hot” + 1 “cold” strand autoradiograph...
MECHANISM OF DNA REPLICATION
Kornberg (late ‘50s)
enzyme that catalyzes DNA replication ... DNA polymerase I
several others
MECHANISM OF DNA REPLICATION
DNA replication requirements... 1. H bonds between bases must be broken 2. chain separation/unwinding 3. available pools of 4 dNTPs: A = T, C G 4. enzymes
E. coli... ~4.7 × 107 bp (47,000,000) molecule has ~ 4.7 × 106 turns (10 bp/turn) 1500 bp/s, ~ 20 min to divide with 1 origin
MECHANISM OF DNA REPLICATION prokaryotes have a single origin of replication bidirectional replication with 2 forks single terminus ~ 20 minutes (*... book says 40)
MECHANISM OF DNA REPLICATION E. coli origin of replication tandem repeat consensus sequence protein binding sites
MECHANISM OF DNA REPLICATION eukaryotes have multiple origins of replication bidirectional replication with 2 forks 1.4 h (yeast) to 24 h (cultured animal cells)... or more issues...
linear vs circular coordination of multiple chromosomes
pulse/chase and autoradiography ...