3.2 Replication Flashcards
Why do we care about replication?
- Transmission of genetic info is essential
- Better understanding of mutations leads to development of better therapeutics, evolution…
Purines are
Adenine and guanine
(Have 2 rings)
Pyrimidines are
Uracil, Thymine, and Cytosine
(Has 1 ring)
Name for DNA sugar
2’-Deoxyribose
(lacks a hydroxyl group at 2’)
Name for RNA sugar
Ribose (RNA)
Chargaff’s rule
A=T
G=C
A-T binding has how many hydrogen bonds?
2
C-G has how many hydrogen bonding?
3 H bonds
Chargaff’s rule was instrumental in helping ___ to decipher the structure of DNA using the X-ray crystallography images of the molecule by ___. The idea to use X-ray crystallography belonged to ___.
Watson and Crick
Rosalind Franklin
Maurice Wilkins
The double helix is ___ with what types of grooves?
Antiparallel
Has major and minor grooves
B-DNA
10.5 bp/turn
Relaxed DNA
The typical DNA
A-DNA
11 bp/turn
(More tightly wound)
Z-DNA
12 bp/turn
Left-handed helix
Most DNA found is nature is ___ supercoiled DNA. It turns in what direction?
Negative
- Turns left
- Less than 10 bp/turn
What is overwound DNA called?
Positively supercoiled DNA
- More than 10 bp/turn
- Turns right
When is positive supercoiling mostly seen?
During replication and transcription
What is nicked DNA?
DNA with a broken phosphodiester bond on 1 strand
- ONLY ON 1 STRAND
Define open circle DNA
circular DNA with a nick
Describe bacterial chromosomes
- They’re circular and negatively supercoiled
- Has DNA binding proteins to compact to form supercoiled domains
(Has histone LIKE proteins, but NO HISTONES)
What is the cytoplasm region with the chromosome called in bacterial chromosomes?
Nucleoid
DNA is eukaryotic chromosomes are bound to equal weight of ___. What do these small basic proteins have large amounts of?
Histones
Arg and Lys
Define chromatin
DNA with protein complex
Define nucleosomes
Beads with protruding DNA (beads of nucleosome cores)
- Composed of 8 histones and 140 base pairs
Define solenoid
Winding of strings of nucleosomes into helical tubular coil
Define heterochromatin
Tightly packed DNA
What is facultative heterochromatin?
Heterochromatin that encodes for genes and can be expressed
- Heterochromatin that is transcribed but constantly turned over via RNA induced silencing
What are the components of eukaryotic chromosomes?
- DNA bound to histones
- Chromatin
- Nucleosomes
- Solenoid
- Heterochromatin
DNA polymerase needs a ___ and can’t synthesize ___
Template
De novo (can only extend the chain)
DNA gyrase works in front of
The replication fork
DNA grows in what direction
5’ to 3’
- DNA pol only works in 1 direction
Define DNA polymerase
Catalyzes elongation of the chain based on the template strand
- Multiple in every organism
How many DNA polymerase’s are in E. coli?
5 DNAP (DNA III = major)
How many DNA polymerase are in humans?
15 (Pol delta and PolE)
Define topoisomerases (Class II)
Relieve stress due to positive supercoiling and reintroduce negative supercoiling
Define helicase
Breaking of H-bonds between bases
Define primase
Synthesis of RNA primers that provide 3’OH for DNAP
What removes RNA primers?
DNAP 1
Flap endonuclease I (FEN1) and RNase H (eukaryotes)
Define DNA ligase
Joining of DNA via phosphodiester bond
Define Proliferating cell nuclear antigens (PCNA)
- Only in eukaryotes
- DNA clamp that enhances DNAP activity at the replication fork
- Diagnostic marker for proliferating cells
- Important cancer diagnostic marker
Most bacteria have ___ chromosomes with 1 ___
Circular chromosomes
1 oriC
oriC is ___rich
AT-rich (meaning A-T)
- Easier to open up
Steps for bacterial DNA replication:
- Formation of pre-priming complex to open up oriC
- SSB bind (protects from zipping back and protects ssDNA)
- Replication fork formation and H bonds break
- Gyrase relieves pos. supercoiling
- Primase
- Bidirectional chain elongation
- Topoisomerase IV (4) reintroduces neg. supercoiling on new DNA
- DNAP 1 removes RNA from Okazaki fragments
- DNA ligase joins fragments
- Chromosome separation
DNA pol I can exonuclease and has activity in what direciton?
5’ to 3’ direction
Leading strand
DNA synthesizes new strand in 5’ to 3’ direction as 1 long strand
Lagging strand
Synthesized in short fragments b/c orientation of template strand is opposite to the direction of DNA synthesis
How many base pairs are okazaki fragments in bacteria and eukaryotes?
~1,000 bp in bacteria
~150 - 200 bp in eukaryotes
Trombone model of DNA replication
DNA of lagging strand loops so it’s in correct orientation for DNA pol
- The leading and lagging strand now contact each other and are replicated at same time
How many RNA primers for leading and lagging strand?
1 for leading and many for lagging
How many termination sites are there in e. coli?
6
Majority of DNA replication occur during what phase?
S phase
Pol__ for leading strand and Pol ___ for lagging strand
Pol E for leading
Poldelta and Polalpha for lagging
Define telomeres
Repeating sequence that may be repeated thousands of times
- Aging associated with shortened telomeres
Define telomerase
RNA dependent DNA polymerase carrying own RNA primer
- Binds to telomere (3’ overhang) and base pairs with it
- Allows for less shrinkage and we can live longer
Define mutations
Heritable changes in genetic makeup that can be beneficial, harmful, or silent
When can mutations be acquired?
During replication or post-replication (damages)
- Many are during
Types of mutations:
Base pair change
Py:Py dimers
Chemical alteration of bases
Insertions
Deletions
Define base pair change
Substitution of one base for another
- Most common
- Ex: A-C instead of A-T
Insertions and deletions cause
Frameshifts
Define mutagens and what are the 3 types?
Agents that cause mutations
Types: chemical, physical, biological
Chemical mutagen examples
ROS (reactive oxygen species), deaminating agents, alkylating agents, base analogs, metals
Example of physical mutagen
UV
Example of biological mutagens
Transposons, plasmids and viruses
What is the proof reading/editing function of DNAP?
DNAP senses mutations and uses 3’ to 5’ exonuclease activity to chew back and remove the wrong base
THEN uses 5’ to 3’ polymerase activity to synthesize the strand
Mismatch repair proteins in e coli
MutH
MutL
MutS
Define mismatch repair proteins
Bind and catalyze removal of a segment of newly synthesized DNA including the mismatch
How does mismatch repair distinguish the parents strand in bacteria?
Methylation at GmATC
- Knows not to nick there
Steps for nucleotide excision repair
- Specific endonucleases cleave the strand with the mutations and it’s removed
- Gap repaired by DNAP
- Ligase repairs the break
Steps for base excision repair:
- removal of altered base by specific N-glycosylase
- AP endonuclease removes sugar and P
- Repair of gap
Define recombination
Breaking and rejoining of DNA molecules in new combination
What are the 2 types of recombination?
Homologous and site-specific (like transposons)
Define homologous recombination
Occurs between identical or very similar sequences and needs extensive region of homology
- Holiday junction
Define transposable elements (transposons)
Genetic elements that can “move” or “hop” around
- They’re important for evolution of organisms
Define transposition and give example
Movement of DNA mediated by transposable elements (transposons)
- Ex: nonhomologous recombination
Define transposase
The enzyme that catalyzes transposition
Describe reverse transcription
RNA to DNA
- catalyzed by reverse transcriptase
- replication strategy of retroviruses
