lecture 3.1: deoxyribonucleic acid · 2020-04-02 · shine laser through a simple spring. laser...
TRANSCRIPT
History DNA synthesis
Unit 3: Molecular Genetics
Lecture 3.1: Deoxyribonucleic Acid
John D. Nagy
BIO 181: General Biology for Majors, Scottsdale Community College
2020 Revision
John Nagy Lec 3.1: DNA 1/37
History DNA synthesis
Outline
1 History and SignificanceHypothesesEvidenceApplications
2 DNA Replication MechanismsEarly ideasModern understanding
John Nagy Lec 3.1: DNA 2/37
History DNA synthesis Hypotheses Evidence Applications
1953 Hypothesis I: DNA Triple Helix
Proc. Nat. Acad. Sci. USA 39:84–97. [2]John Nagy Lec 3.1: DNA 3/37
History DNA synthesis Hypotheses Evidence Applications
1953 Hypothesis II: DNA Double Helix
James Watson and Francis Crick Rosalind Franklin Maurice Wilkins
Nature (1953) 171:737–738. [3]
John Nagy Lec 3.1: DNA 4/37
History DNA synthesis Hypotheses Evidence Applications
Visualizing a molecule without microscopes
X-ray crystalography
1 Crystalize the sample (remove allthe water).
2 Shine X-rays through the crystal.
3 Image the diffraction pattern(sort of like a shadow).
4 Use the pattern to reconstructelectron densities.
5 Use the reconstruction toestimate molecular structure.
John Nagy Lec 3.1: DNA 5/37
History DNA synthesis Hypotheses Evidence Applications
Example
Shine laser through asimple spring.
Laser acts like x-rays;spring acts like themolecular sample.
Note that the twisted,helical spring producesan X-shapeddiffraction pattern.
Also note that detailsof the diffractionpattern are directlyrelated to exact shapeof the spring, like howtightly coilded it is.
John Nagy Lec 3.1: DNA 6/37
History DNA synthesis Hypotheses Evidence Applications
Franklin’s X-ray Crystalograph of DNA
Compare to the last example. Does this pattern support orcontradict either or both DNA shape hypotheses?
John Nagy Lec 3.1: DNA 7/37
History DNA synthesis Hypotheses Evidence Applications
The Watson-Crick-Franklin-Wilkins model
John Nagy Lec 3.1: DNA 8/37
History DNA synthesis Hypotheses Evidence Applications
Consequences of discovery: DNA Sequencing
John Nagy Lec 3.1: DNA 9/37
History DNA synthesis Hypotheses Evidence Applications
Consequences of discovery: Human Genome Project
Source: www.ncbi.nlm.nih.gov/projects/genome/guide/human
John Nagy Lec 3.1: DNA 10/37
History DNA synthesis Hypotheses Evidence Applications
Consequences of discovery: PCR
John Nagy Lec 3.1: DNA 11/37
History DNA synthesis Hypotheses Evidence Applications
Consequences of discovery: Molecular surgery
John Nagy Lec 3.1: DNA 12/37
History DNA synthesis Early ideas Modern understanding
A momentous observation
Watson and Crick (1953)
“It has not escaped ournotice that the specificpairing we havepostulated immediatelysuggests a possilbecopying mechanism forthe genetic material.”–Nature 224: 471.
John Nagy Lec 3.1: DNA 13/37
History DNA synthesis Early ideas Modern understanding
Here’s what they meant
See also: dnalc.cshl.edu/resources/3d/01-replication-the-helix.html
But this simple view is wrong-ish.
John Nagy Lec 3.1: DNA 14/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
15’3’
5’
3’
DNA ReplicationSlide number: 1
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Helicase binds to origin and separates strands.
1
John Nagy Lec 3.1: DNA 15/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
2
5’3’
5’
3’
Binding proteins prevent single strands from rejoining.
2
Helicase binds to origin and separates strands.
1
DNA ReplicationSlide number: 2
John Nagy Lec 3.1: DNA 16/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
3
3’5’
5’3’
5’
3’
Primase makes a short stretch of RNA on the DNA template.
3
Helicase binds to origin and separates strands.
1
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 3
John Nagy Lec 3.1: DNA 17/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
3
3’5’
5’3’
5’3’
5’
3’
Primase makes a short stretch of RNA on the DNA template.
3
Helicase binds to origin and separates strands.
1
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 4
John Nagy Lec 3.1: DNA 18/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
Overall directionof replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
4
5’3’
5’3’
5’
3’
3’5’
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 5
John Nagy Lec 3.1: DNA 19/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’3’Overall directionof replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
45’3’
5’
3’
3’5’
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 6
John Nagy Lec 3.1: DNA 20/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
4
5’3’
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Overall directionof replication
3’5’
5’
3’
3’
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 7
John Nagy Lec 3.1: DNA 21/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
4
5’3’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Overall directionof replication
5’
3’
5’3’
3’
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 8
John Nagy Lec 3.1: DNA 22/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5
5’3’
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Overall directionof replication
5’3’
5’
3’
3’
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
Binding proteins prevent single strands from rejoining.
2
DNA ReplicationSlide number: 9
John Nagy Lec 3.1: DNA 23/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
6
5’3’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’3’
3’5’
3’Overall directionof replication
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 10
John Nagy Lec 3.1: DNA 24/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’3’
7
3’5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Overall directionof replication
Okazakifragment
5’
5’3’
3’5’
3’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 11
John Nagy Lec 3.1: DNA 25/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’3’
7
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
Overall directionof replication
Okazakifragment
5’3’
3’5’
3’
5’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
3’
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 12
John Nagy Lec 3.1: DNA 26/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’ 3’5’
3’
3’
5’3’
3’
5’ 5’3’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 13
John Nagy Lec 3.1: DNA 27/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’3’
3’5’
3’5’3’
5’ 5’3’3’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 14
John Nagy Lec 3.1: DNA 28/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’3’
3’5’
3’3’
’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 15
DNA polymerase I remove RNA primers, replaces them with DNA.8
John Nagy Lec 3.1: DNA 29/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
5’
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’3’
3’5’
3’3’
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Binding proteins prevent single strands from rejoining.
2
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 16
DNA polymerase I remove RNA primers, replaces them with DNA.8
John Nagy Lec 3.1: DNA 30/37
History DNA synthesis Early ideas Modern understanding
Conceptual schematic of DNA replication
Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.
5’
3’5’
3’
5’
3’ 5’
3’
Helicase binds to origin and separates strands.
1
Primase makes a short stretch of RNA on the DNA template.
3
DNA polymerase proofreadingactivity checks and replacesincorrect bases just added.
5
Leading (continuous) strand synthesis continues in a 5’ to 3’ direction.
6
Discontinuous synthesis produces Okazaki fragments on the 5’ to 3’ template.
7
Binding proteins prevent single strands from rejoining.
2
9 DNA ligase repairs nick in sugar phsophate backbone.
DNA polymerase I remove RNA primers, replaces them with DNA.8
DNA polymerase III adds DNA nucleotides to the RNA primer.
4
DNA ReplicationSlide number: 17
John Nagy Lec 3.1: DNA 31/37
History DNA synthesis Early ideas Modern understanding
Details of DNA polymerase III action
Catalytic steps of elongation by DNA polymerase III1 Polymerase “reads” the proximal unpaired nucleotide on the template
strand.
2 It aligns the complementary nucleotide triphosphate; catalyzes formation ofhydrogen bonds.
3 Cleaves β and γ phosphates from nucleotide triphosphate; uses the releasedenergy to form the phosphodiester bond between added nucleotide andgrowing DNA strand.
4 Slides one nucleotide on the template strand and repeats.
John Nagy Lec 3.1: DNA 32/37
History DNA synthesis Early ideas Modern understanding
Details of elongation
John Nagy Lec 3.1: DNA 33/37
History DNA synthesis Early ideas Modern understanding
DNA synthesis schematic: Chromosome ends
John Nagy Lec 3.1: DNA 34/37
History DNA synthesis Early ideas Modern understanding
A more realistic picture
Source: [1]
Definition: replisome
This replisome is a complex of the enzymes of replication. Seealso:dnalc.cshl.edu/resources/3d/04-mechanism-of-replication-advanced.html
John Nagy Lec 3.1: DNA 35/37
History DNA synthesis Early ideas Modern understanding
Summary
When studying this process, organize your thoughts aroundthese concepts:
Phases:InitiationElongationTermination at chromosome ends
Enzymes involved:Helicase and topoisomeraseSingle Strand DNA-binding proteins (SSBPs)PrimaseDNA polymerase IIIDNA polymerase ILigase
Leading strand and lagging strand
John Nagy Lec 3.1: DNA 36/37
History DNA synthesis Early ideas Modern understanding
References I
Jong-Bong Lee, Richard K. Hite, Samir M. Hamdan, X. Sunney Xie, Charles C.
Richardson, and Antoine M. van Oijen.DNA primase acts as a molecular brake in DNA replication.Nature, 439:621–624, 2006.
Linus Pauling and Robert B. Corey.
A proposed structure for the nucleic acids.Proc. Nat. Acad. Sci. USA, 39:84–97, 1953.
James D. Watson and Francis H. C. Crick.
The molecular structure of nucleic acids: A structure for deoxyribose nucleic acid.Nature, 171:737–738, 1953.
John Nagy Lec 3.1: DNA 37/37