final Flashcards
what is chromatin a mixture of
dna and proteins that form chromosomes
what do the proteins in chromatin do, what is the main one
they pack dna into a compact form, especially histones
what phase of the cell cycle is dna especially compact
metaphase
what is chromosome structure during each phase of cell cycle
interphase : chromosomes are present as chromatin (uncondensed)
s phase (DNA synthesis): each chromosome is replicated, resulting in 2 sister chromatids
prophase: chromosome becomes more condensed
what happens to chromosome once cell division is complete
becomes less condensed
what is euchromatin
less condensed form of chromatin, in non-dividing cells
what is heterochromatin
more condensed form, in diving cells
what do histones do
protect and package the dna and regulate replication and transcription by controlling access to the machinery
what is a nucleosome
dna wrapped around histones
fundamental unit of chromatin (beads on a string)
histones packages and orders dna into them
what does a core histone comprise of
2 copies of each, form an octet (8 subunits)
what chromatin is 11 nm and what one is 30nm
11 - beads on a string - heterochromatin
30- solenoid - euchromatin
what holds dna on histone core
H1 histone
how much h1 is there on genes being actively transcribed (being used as templates)
little to none
what is higher order chromatin structure attached to
nuclear/chromosome scaffold
what do histones mainly react with , what does this explain
phosphate backbone on dna
explains their lack of specificity
what mediates assembly of 30nm fiber
N terminal histone tail
what is 30nm finer held together by
h1 proteins - pull nucleosomes together
N terminal tails - bind dna on existing nucleosomes
does gene transcription occur in solenoid
no
what is chromatin remolded by
enzymes that covalently modify histones (ex HATs)
chromatin remolding complexes (increase decrease accessibility to dna
what 3 dna sequences control separation and copying of chromosomes
replication of origins
centromere
telomere
what is the replication origin
where dna duplication begins
why do eukaryotic chromosomes contain may replication origins
to ensure they can be duplicated rapidly
what is a centromere
attachment site for biotic spindle via protein complex called kinetochore
what is a telomere
formed at end of chromosomes
like aglet on shoelaces
protects end of chromosomes from shortening with each cell division
highly repetitive - so not recognized as dna
what do telomerases have as part of their active site and why
RNA molecule
acts as template for extension of telomere, which allows their degradation
why are telomerases a reverse transcriptase
since they use ran template to make dna
why are the 3’ ends of chromosome ssDNA
degradation of rna primer
what happens to 3’ ssDNA
dsDNA is synthesized from extended 3’ ends using ran primer, dna pol
remaining ss 3’ oh ends are protected by looping and dna binding proteins
telomeres
what is primary structure
nucleotide sequence
what is secondary structure
stable structure adopted by a segment of dna
typically a base paired double helix
what is a tertiary structure
3d fold
complex folding into super coils and chromatins
what are the purines
adenine
guanine
what are the pyridines
thymine, uracil, cytosine
what does base + sugar =
nucleoside
what is base + sugar + phosphate =
nucleotide
what is pentose in rna and dna
ribose in rna
deoxyribose in dna
what does rna have on 2’ carbon
2’-OH
what does dna have in 2’
2’-H (acid)
is the 2’-oh of rna or 2’-h of dna less stable
rna is less stable and more reactive
where is ribose attached to base on nucleoside
attached to base via its 1’ carbon
what attacks phosphate
5’ c of pentose
what is 3’-oh of one nucleotide is linked to 5’ phosphate on adjacent nucleotide forming ?
sugar-phosphate backbone of DNA
how does phosphate group link pentoses
phosphodiester bond
what happens to phosphate group of every nucleotide at physiological pH
its deprotonated so they carry a negative charge
how many h bonds are between G and C
3
how many h bonds are between A and T/U
2
what type of linkage is the bond between pentose and base
beta linkage
what does a b linkage mean
base lies in plain above the sugar
what does an alpha linkage mean
base lies below the plane of sugar
how do ribonucleotides differ from deoxyribonucleotides
have hydroxyl at the ribose 2’ (not deoxy)
uracil
what holds two dna strands together
h bonds
do dna strands run in same direction
no, they’re anti parallel
in addition to h - bonds what else holds double helix together
base stacking between aromatic hydrophobic bases
what does base stacking do
minimizes contact with h2o, stabilizing it
what force is base stacking a form of
van der waals
what does base stacking result in
minor / major grooves
what are the difference in depth of major and minor grooves
major = deep
minor = shallow
what type of groove are base pairs more exposed to solvent
major
where are functional groups in base pairs that aren’t involved in bp more exposed, and what is the purpose of this
in grooves, especially major
these exocyclic groups serve as contact points for proteins
what is a b form (b-helix/bDNA)
form normally found in cells
right handed
most stable under physiological conditions
what is a-form of dna
when dna is dehydrated it assesses the a form
right handed double helix
wider shorter, bp are more tilted, deeper major grooves, shallower minor grooves
where is a form frequently found
dna-rna hydrids
ds RNA (tRNA)
what is z-form dna
left handed and stretched
what frequency do nucleotide bases absorb light at
260 nm
what is TM
melting point
temperature at which half of dna is ss and other half is ds
do stacked bases absorb more or less UV light
less
hypochromism
what determines what temperature TM occurs at
nucleotide sequence length
[salt]
what does high [salt] and longer sequences result in, why
stablize sequence, increase TM
salt ions shield negatively charged phosphates on DNA backbone, which repel each other when unshielded
under what circumstances can supercoiling only occur
in a molecule where both strands are closed circles or fixed at one end
what a LK
of times one strand crosses over the other
LK for closed circular DNA is always an integer
what level of wound is most cellular dna
umderwound (fewer helical turns than expected for B form)
what does unwinding induce and what does it do to accommodate
structural strain
can accommodate by supercoiling or break H bonds (usually super coiling)
what does cells underwinding their dna using topoisomerases to result in a strained state result in
form of stored energy
what does underwound supercoiled state allow
compact packaging of DNA
what is the only way LK can change
unless one or both of the strands is broken
can be changed by means of topoisomerases
does super coiled dna more slower/fatser
faster
what is a topoisomerases
enzymes that catalyze interconversion of topoisomers (change supercoiling by cutting / splicing DNA)
chang LK
how do topoisomerases change the LK
- cleavage of one/both strands of DNA
- passage of segments of DNA through this break
- resealing of DNA breaks
what are the two types pf topoisomerases
TOPO I : change LK by increments of 1
TOPO II : removing 2 turns at a time
what is protein synthesis
translation
nucleotide sequence in mRNA is translated by tRNAs into amino acid sequence in polypeptides
what is link between DNA and protein
RNA
where do steps of protein synthesis take place in eukaryotes
DNA is confined to the nucleus, RNA is made in nucleus (transcriptions) but is exported to cytoplasm
protein is synthesized in cytoplasm
what is protein synthesis proportional to
amount of RNA in cell
what is mRNA
intermediary between gene and protein
provides transcript that is read by tRNA
what is tRNA
reads genetic code, brings amino acids to the growing polypeptide chain
what is rRNA
in ribosome
provide scaffold for protein synthesis, catalyze peptide bond formation
do amino acids bind directly to mRNA
no
what is a frameshift
insertion or deletion of one of two bases result in frameshift
how is nucleotide sequence in mRNA translated into an amino acid sequence in proteins
- enzyme called tRNA synthase attaches an amino acid onto one end of tRNA
- tRNA brings that amino acid to growing polypeptide chain in the ribosome, selected by base pairing between 3’ nucleotide and anticodon on the tRNA and the 3 nucleotide codon on mRNA
are tRNA and mRNA parallel
no
what is a start codon
at start of gene - signals start of protein synthesis
what is a stop codon
signal the end of gene
terminate protein synthesis
do stop codons encode an amino acid
no
what direction does tRNA read mRNA
5’ to 3’
what are the 4 steps of polypeptide synthesis
- initiation
- elongation
- termination
- ribosome recycling
what are ribosomes
ribonucleoproteins that catalyze protein translation
what are the 3 sites on a ribosome
a site : for aminoacyl tRNA
P site : for peptidyl tRNA
E site: for exit
what direction do ribosomes tracks along the mRNA
5’ to 3’
N to C
what are the steps of initiation
- GTP and initiation factors bind to 30S ribosomal subunit
- complex tracks along mRNA until it finds the ribosomal binding site (RBS) (In banteria its the shine-dalgarno sequence SDS), positioning the the start codon in the P site. then a special initiation tRNA charged with a modified methionine (fmet) binds to that start codon
- the 50S ribosomla subunit binds, GTP is hydrolyzed and initiation factors are released, forming a functional 70s subunit initiation complex , with modified methionine (fmet) in P site, with A and E site empty
what are the steps of prokaryotic initiation
- initiation factors bind to 30s subunit. some bind to/hydrolyze GTP to facilitate steps in translation
- since SDS is upstream of start codon on the mRNA, it positions the start codon on the P-site. 30s subunit recognizes SDS via bp and rRNA
- Initiation factor is released and the 50S subunit binds to the 30S
GTP is hydrolyzed, facilitating release of ignition factors, resulting in 70s ignition complex
what is the only thing that can bind to P site
Initiating tRNA (fmet - tRNA ) can bind to p site
all other amino acids bind to a site
why is GTP like ATP
has high energy phosphodiester bonds
hydrolysis of these bonds provide energy
what are the steps of elongation
- elongation factor binds to GTP and then to an amino acid - tRNA
- GTP binds to 70s initiation complex at the a site via base pairing with mRNA
- GTP is hydolyzed and GTP is released and recycled
- the rna in the 50s subunit catalyzes the transfer of amino acid #1 (fmet tRNA) to the new amino acid, forming new peptide bond
- for the ribosome to translocate one codon along the mRNA to next site of amino acid addition, a second elongation factor binds to GTP and associates with the A site
- GTP is hydrolyzed and ribosome is translocated along mRNA in 5’ to 3’ direction. new initiating tRNA is in P site, and discharged tRNA is now in e site where its released, opening a site
what catalyzes the transfer of growing polypeptide to from tRNA in p site to the amino acid on the rna in the a site
ribozyme activity of rRNA within the p site of 50s subunit
during peptidyl transfer what acts as nucleophile
amino n of incoming A site amino acid , attacking C electrophile in 3’ terminus of p site
what is an termination/release factor (RF)
protein that mimics tRNA transfer and can bind to a site when a stop codon is present
what happens in bacteria when a codon occupies a site
a GTP bound RF binds to a site, which causes dissociation of 70s subunit
what initiates protein synthesis of eukaryotes
mRNA binds to ribosomes at 3’ poly A tail by poly A binding protein (PABP)
what is the only thing that can bind to the small subunit at p site without complete ribosome present
only the met charged with initiating tRNA met can bind to it
what is initiation powered by
ATP and GTP hydrolysis
what do chaperons do
prevent proteins from aggregating via exposed hydrophobic regions
shield hydrophobic regions
also help fold misfolded proteins correctly
what destroys misfolded proteins
atp- dependent protease complex called proteasome
what does ubiquitin do
marks proteins with exposed hydrophobic regions for destruction
allows them to be recognized by proteasome
what is a mobile genetic element
nucleic acid segments (DNA/RNA) that can move in/out of cells or between different regions of of cell genome
what is recombination
when some mobile genetic elements get integrated into host genome
what are the types of mobile genetic elements
plasmids
viruses
transposable elements (transposons)
free dna
what is a plasmid
extra chromosomal circular ds dna found in bacteria, yeast, fungi
replicates with host machinery
lack protein coat and cant more independently from cell to cell, can through conjugation
what is a virus
infectious dna or rna containing elements that possess a protein coat that allows them to move from cell to cell
replicate using host proteins, use some of their own which are synthesized into host
what are transposable elements
lack a protein coat and can be excised from one part of host genome and can insert into another part via recombination
what is free dna
from lysed bacteria, can be taken up by other related bacteria
what do plasmids often encode
functions that can give an advantage to host such as abiotic resistance
do plasmids tend to incorporate into host genome
no
what are virulence factors
may contribute to virulence (severity) of pathogen
what are plasmids related to
bacteriophage genomes
what happens to lytic phage genome
doesn’t integrate into the host chromosome, replicated extra-chromosally
assembled phage often lyse their host cell when they’re released
what is lysogenic phage genome
becomes integrated into host genome via recombination , forms a prophage that is replicated along with host dna, passed onto daughter cells
how can plasmids, phage, and free dna can be passed from one cell to another
horizontal gene transfer :
- transformation: uptake of naked DNA
- transduction: injection of foreign dna by a bacteriophage
- conjugation: compatible strains join via a conjugative pilus - conjugation bridge forms
what do restriction endonucleases do
degrade viral genome
recognize a short DNA sequence (a restriction site) and cut the strands
what produces restriction endonucleases (RE)
by bacteria to protect from viral infection
what does a polymerase chain reaction (PCR) do
amplifies specific DNA sequence
so you don’t need to cut gene of interest out
when must rna bp with dna
transcription
is rRNA coding
no
in prokaryotes what are all RNAs transcribed by
a single RNA polymerase
where does bacterial transcription take place
all in cytoplasm (no nucleus)
what processes are coupled in bacteria
transcription + translation
both occur in same direction (5’ to 3”) and everything is synthesized in cytoplasm so protein synthesis can occur as mRNA is being synthesized
what regulates protein expression in bacteria
mRNA production
what does [mRNA] depend on
rate of synthesis/degradation of mRNA
what are three elements of gene control in prokaryotes in order of importance
- transcription initiation
- rna turnover (by nuclease degradation)
- transcription termination
what does dna transcription produce
ss rna
what does prokaryotic transcription require
rna polymerase
ds dna template
ribonuclease (NTP)
Mg 2+
no primer !
what do Mg ions do
coordinate the phosphates of incoming NTPs, position P alpha for nuc attack
what do ribonucleases (NTP) do
breaks down the RNA by cleaving the phosphorus-oxygen bonds.May 20, 2022
what does RNA pol do to initiate rna syntheis
bind to promoter sequences,
recognizes promoter via sigma factor
what does RNA pol + sigma subunit =
rna polymerase holoenzyme
what is the bacterial transcription cycle
- initatition: rna polymerase holoenzyme forms, binds non-specifically to DNA
- polymerase partially unwinds DNA (has helices like activity), opens complex and begins transcribing
- after about ten ribosucleitides are polymerized, polymerase undergoes conformation change, releasing sigma factor , enabling it to move forward rapidly
- elongation
- termination
what stage in prokaryote transcription relies on a sigma factor
initiation
where does prokaryotic transcription have to start
at promoter
what is rho
a termination factor
prokaryotes are either rho dependent or independent
what are activators
increase expression levels
what are repressors
decrease expression levels
how do repressors work
bind near promoter and block RNA polymerase transcription
how do activators work
bind near promoter and enhance ability of RNA polymerase
often near weak promoter
what are inducers:
ligand that decrease affinity of the repressor
what are co-repressors
ligands that increase the affinity of repressor
where is transcription machinery in eukaryotes
nucleus
are eukaryotes cells organized into operons
how are they organized
no
transcribed as monocistronic messages with a promoter for every gene
what does rna polymerase I (RNAPI) synthesize
synthesis of rRNA
what does rna polymerase II (RNAPII) synthesize
mRNA
what does rna polymerase I (RNAPIII) synthesize
tRNA
what do eukaryotic polymerases require to bind DNA
transcriptions factors
in eukaryotes what provides transcription for a given promoter
TF
what does RNAP I transcribe
transcends 1 gene
4ss precursor rna for rRNA
how much stuff does RNAP II transcribe
tens of thousands
how much stuff does RNAP III transcribe
hundreds
what does binding of RNAP II factors to promoter do
imitates polymerization
do eukaryotes use positive or negative regulation more
positive
where do activators bind in eukaryotes
specifically to enhancer sequences near promoter
how do activators help assemble polymerase initiation complex
by recruiting RNAP II
what mediates splicing
spliceosome complex that associates with RNAP II
what are introns
non coding regions
what is the difference between group 1 and 2 introns
The key difference between group I and group II introns is that in group I introns, the splicing reaction is initiated by a guanosine cofactor, while in group II introns, the splicing reaction is initiated by internal adenosine
what is dna replication
duplication of chromosomes
what are the dna building blocks
deoxynucleoside monophosphate (dNMP or NMP)
what does replication occur by
addition of NMPs to 3’-OH of a nascent (new) strand
what direction does dna synthesis occur in
5’ to 3’
what are the raw materials of dna replication
dna template
deoxynucleoside triphosphate (dNTPs)
Mg2+ ions (cofactor for polymerase)
DNA/RNA primer to provide first 3’-oh
dna polymerase
what is DNA polymerase
large protein complex that includes the enzyme that catalyzes the addition on NMPs to DNA polymer
what do DNA polymerase reactions add to the 3’ end of growing strand
deoxynucleotide
what does Mg2+ do
stabilize negative charges on incoming dNTP and assists in deprotonation of 3’-OH
what does DNA pol III do
main polymerizing enzyme
what does DNA pol I do
completes Okazaki fragment synthesis (replace RNA with DNA)
what direction is polymerization always in
5’ to 3’
what is exonuclease
an enzyme that catalyzes the hydrolysis of phosphodiester bonds at the end of nucleic acid molecules (cuts DNA strand)
where does dna replication occur
replication forks
what are the pimosomes and what do they do
primase: generates rna primer
helicase: unwinds dna
why does lagging stand loop around
allows both dna pol III subunits to move together in same direction
what is processivity
the ability of an enzyme to repetitively continue in catalytic function without dissociating from substrate
what dna pol is less processive, why
dna pol I
doesn’t have clamp
what does the beta clamp allow for and not allow for
allows for processivity of leading but not lagging strand
what is nick translation
process of simultaneously adding NMPs to the 3’ end of Okazaki fragment and removing NMPs from the adjacent from the adjacent 5’ end of the rna primers
completes synthesis of lagging strand
what do incorrect bases result in
steric clashes
