Ch 6 Exam 2 Flashcards
What did the meselson-Stahl experiments show
That DNA replication is semiconservative
What’s the really important enzyme that DNA replication needs
DNA polymerase. 5’-3’ prime
WHat is released when a nucleotide is added by DNA polymerase
a pyrophosphate
What are the lagging strands called
oakizaki fragments
what are the four steps of lagging strands
primer synth
elogation
primer removal with gap filling
joining of okazaki fragments
What is a replisom
a multiprotein machine with a suite of basic enzmatic functions:
Helicase
primase
dna polymerase
3’ to 5’ exonuclease
WHat are single strand binding proteins
stabling single strand so it doesn’t fold on it’s self
RPA in humas
SSB in bacteria
What does DNA polymerase do
strand elongation
RNA primer removal
Which polyimerases are for replication of
1 and 3
DNA polymerase 1
Primer removal, gap filling between okazaki frgaments and nucleotide excision repair pathway
Two fragments of pol 1
Klenow fragment has 5’ to 3’ polymerase activity and 3 to 5 exonuclease activity(for proof reading)
DNA polymerase III
main replicative polymerase. both 3 to 5 and 5 to 3
DNA polymerase II, IV, V
DNA repair mechanisms
Where does dna rep begin
origin.
some have as many as 3
A-T rich
DnaA can only bind to negatively supercoided origin DNA
WHat does helicase do
unwind
Steps in rep innitiation
Initiator protein (DnaA) binds to Origin
DNA helicase unwinds helix
two replication forks are formed
replication is bidirectional
Two sliding clamps tether DNA polymerase to DNA polymerase
Single-standed binding proteins (SSBPs) keep dna helix open and protec from nuclease attack
Primase synthesize RNA primer
WHat’s a clamp loader
uses ATP to open and close the sliding clamps aroud DNA
steps for DNA elongation
two molecules of DNA polymerase III catalyzes DNA synthesis at the time.
DNA template
all four deoxyribonucleoside triphosphates (dNTPs)
primer
what’s the main polymerase for elongation
polymerase III
What direction does DNA polymerase synthesize and what direction does it read
5’ to 3’
reads 3’ to 5’
what removes the RNA primer and syntehsize DNA complementary deoxyribonucleotdies
Poly 1
WHat does DNA ligase do
catalyzes phosphodiester linkage between adjacent fragments of. seals it up
what’s the end site of replication
Ter site
What topoisomers
forms of DNA that have the same sequencing but different linking number and mobility. varying amount of coiling
Bacterial type 1 topoisomerase
relax negative supercoils
relax negative and positive supercoiling
can detangle
do not require ATP
single strand breaks
Type II topoisomerase
relax both negative and positive supercoils
unknot or decatenate entangled DNA molecule
Needs ATP^^
gyrase introduces negative supercoid
double sttrand breaks
What can block leading strand synthesis
damaged of template
What are the origin on eurkaroites
internal sites on linear chromosomes
Autonomous replicating sequence
in saccharomyces cerevisiae
origins “fire” how many times per cell division
once
are the origins activated at the same or different times
uniformly activated
E First step
Histone removal.
Histone modification and chromatin remodeling factors are some ideas
What stage does origin selection occur in
G1 Phase
What phase does DNA replication happen in
S
What does the prereplication complex (pre-RC) do
Forms at origin.
Assembly of proteins: ORC (origin recognition complex), Cdc6 and Cdt1 (load MCM helicase complex), and MCM helicase (unwinds the DNA)
Licensing for replication (only happens once)
Transitions from licensed state into S phase to active replication
What is ORC
DNA binding complex that binds Cdc6 to origins
What happens after the Mcm 2-7/Cdt1 binds to Cdc6
Cdc45/GINScomes in and Cdc 6 and Cdt1 leave. After the pre-replication complex is formed (G1) and the cell enter S phase, Cdc45 binds to MCM helicase
What happens after Cdc6 is bound to DNA
Recruits Mcm 2-7 with Cdt1
E What is licensing
It’s a process that ensures that replication only occurs once per cell cycle
Prepares origin to be used during the S phase
What is licensing controlled by
cyclin-dependent kinases (CDKs)
What are CDKs activated by
cyclin
What phase is CDK low and what does that allow
G1, allows Mcm 2-7 can be loaded
CDK is high
S and G2
CDK plumets during M phase causing licensed origins fire, but can’t be reloaded, phosphorylates Mcm2-7. Mcm2-7 can not be loaded
In Eurkaryotes, which strand is the Helicase loaded onto
Leading
in bacteria where does the helicase load
lagging
what are the singlge strand binding proteins for eurkaryotes
RPAs
What’s the RNA primer for eurkaryotes
it is synthesized by DNA polalpha and it’s associated primase
What does pol alpha/primase enzyme synthesize first
7-10m bases of RNA, then 25-35 bases of initiator DNA (iDNA
WHat polymerases are after pol alpha
Leading strand: polymerase e
lagging strand: polymerase delta
handoff between dna pol alpha to the appropriate one
What are the DNA polymerases in mammals used for chromosomal DNA replication?
DNA pol alpha, pol delta, pol epsilon
Mitochon: Pol y
ALl the rest of them: repair
What’s the name of the E clamp
PCNA
loader is RFC. opens and closes clamp, subunits
Steps in proofreading
pause
melting
translocation of incorrect to the exonuclease
removal of the 3’ terminal base
It’s identified by abnormal geometry of mismatcehd base pairs and base-base hydrogen bonding
After repair what needs to happen to
RNA primer removal
gap fill-iin
joining of okazaki fragments on the lagging strand
what fills in and joins okazaki fragments
FEN1 and DNA ligase 1
Histone reassembly
quick.
Rolling circle replication
a common mode of viral and factor replication
Steps in rolling circle replication
donor cell attackes to recipient with pilus
cells contact one another
one strand of plasmid DNA transfers to recipient
recipient synthesizes a complementary strand
What are telomers
end of chromosome and DNA
How to telomerases solve the lagging strand problem?
has a RNP complex that reverse transcriptase activity. . Essentially has a primer to it.
contains an RNA component that provideas a template for telomere repeat synthesis
Two parts of telomer
RNA: Telomerase RNA component (TERC)
Protein: Telomerase reverse transcriptase (TERT)
What happens after telomerase elongates the DNA
it forms a T-loop
WHat helps make the DNA legnth after the telomermase just right
TZAP
Is telomerase always on and being used
yes
Does progressive shortening of telomeres in human somatic cells happen?
yes
What happens when cells have little telomerase activity
Hayflick limit. The cells become senescent. Don’t grow and divide.
shelterin
The telomeres are essential for maintaining chromosomal stability, and shelterin plays a key role in ensuring that they are protected from unwanted DNA damage responses and inappropriate DNA repair processes.
Which strand does telomeres elongate
The 3’ strand ends up longer, no problem, shelterin makes it all safe and good
