Exam 2-My Q Flashcards
Griffith Experiment
experiment on pnemonia injected into mice, found that bacteria can transform to have capsule.
Explain Avery and McCarty experiment
extracted cell contents of bacteria and found the enzyme that inactivated the “transforming principle”. found that the transforming principle was made of DNA.
Explain the Hershey and Chase experiment
infected the bacterial culture with radioactively labelled phages. in P to identify DNA and with S to identify proteins. found that next generation of bacteria still had the radioactive phosphorus, showing that DNA carries information from one gen to the next
Explain what Watson and Crick discovered
DNA structure, in 1956 published that DNA is made of a nitrogenous base, phosphate group and 5 carbon sugar. They also discovered it exists in a double helix
What kind of bonds hold together a DNA mol?
phosphate group-pentose sugar-nitrogenous base.
all covalent bonds.
chain held together by phosphodiester bonds
nucleotides by H bonds
where does the nitrogenous base connect to the pentose and where does the phosphate group connect to it?
nt-c1
p-c5
name the 4 nitrogenous bases and how they are categorized
pyrimidines-Cytosine, Uracil, Thymine
purines- guanine and adenine
Explain Chargaff’s rules (3) and how they were discovered
- A-T and G-C “proportional to”
- The sum of purines is equal to the sum of pyrimidines (A+G)=(C+T)
- The ratio of G+C to A+T varies greatly between organisms
How: through chromatography data
explain what Rosalind Franklin discovered
through x ray diffraction she found data that showed DNA might be some form of helix
Describe the double helix shape of DNA
2 antiparallel polynucleotide chains
coiled around a central axis
twisted into a right handed helix
What are the chains in the double helix made of and how are they bonded, describe the inside bonds as well
a sequence of covalently bonded nucleotides with a phosphate sugar backbone , with the base pairs located on the inside of the helix
base pairs bonded by H
backbone bonded by phosphodiester bonds (covalent bond)
Describe what is at the 3’ and 5’ end of the DNA chains
3’-free 3’C with an OH (hydroxyl)
5’-free 5’C with a phosphate
name nucleotide chains with:
2 nucleotides
less than 20 nucleotides
more than 20 nucleotides
2: dinucleotide
<20: oligonucleotides
>20: polynucleotide
explain what antiparallel means
opposite orientations with regards to the 3’ and 5’ end
What kind of bonds hold together the base pairs
G-C triple H bond
A-T double H bond
What are nucleosomes made of
DNA wrapped around histone octamer (a protein). allows length of DNA to shrink 1/3
describe solenoid
coiled and stacked nucleosomes
describe chromatin fibers
solenoids that are folded with looped domains
describe a chromosome.
chromatin fibers that are compacted into x structure that is only visible during cell division.
Describe the structure of eukaryotic chromosomes in order of smallest to largest scale
nucleosomes, coiled and stacked into solenoids, folded into chromatin fibers, looped into the shape of a chromosome.
how many genes are in 1 chromosome
ranges from a couple hundred to several thousand
what are the parts that make up eukaryotic chromosomes
telomere, origins of replication, centromere
how often is there an origin of replication in a single chromosome
about every 100,000 base pairs
what is the purpose of the centromere
allows the chromosome to segregate into daughter cells
What do telomers do?
protect the ends of the chromosome
what are heterochromatin and euchromatin
hetero-never uncoils, remains condensed even during interphase
eu-loosely condensed, becomes uncoiled during interphase, contains most protein coding genes
where are heterochromatin and euchromatin located
eu-on arms of chromosome
hetero-at the centromere, telomere and other places throughout the chromosome.
what are the sequences of euchromatin and heterochromatin like?
eu- unique sequences
hetero- repeated
what can be said about the sequences of eukaryotic genomes?
highly repetitive DNA, made of more introns than exons
what is semiconservative, conservative and dispersive DNA replication, which theory is correct? who proposed it
Watson and Crick proposed the 3 possible structures
semi-strands of DNA have both new and old DNA,
conservative- new with new, old with old
dispersive-each stand is made of a mix of both old and new
Meselson-Stahl found DNA replication is semiconservative
What are the features of DNA polymerase
- it can synthesize a chain of nucleotides only if there is a primer
- only works with the 3’ end of a growing chain, moves from 3’->5’
What direction does DNA replication occur in
5’–>3’
Explain how DNA replication originate in bacteria?
- DNA strand is circular
- there is a single origin, (oriC)
- replication continues bidirectionally from oriC
- 2 replication forks form and migrate apart
- the forks merge at a terminator sequence (ter)
- result is 2 separate circular DNA strands.
Explain how DNA replication originates in eukaryotes
- DNA is linear and longer
- there are multiple origins of replication on a single chromosome
- replication is bidirectional
- forks either reach the end of the chromosome or run into each other
- end result is 2 sister chromatids
Is DNA replication in eukaryotes continuous or discontinuous? explain
both, it is continuous for the leading strand, which goes in the direction of the fork
it is discontinuous for the lagging strand, which moves away from the fork.
explain what okazaki fragments are
they form in the lagging strand, because the replication is discontinuous separate fragments are formed with gaps between them.
Explain the trombone model
it is the structure that DNA polymerase has when moving along the DNA strand. 2 circular pieces move along the strand, one for the leading strand and one for the lagging. The lagging strand has a loop in order to translate in the 5’ to 3’ direction.
how many different DNA polymerase enzymes do bacteria and eukaryotes have, name the important ones
bacteria- 5, poly III does 5’-3’ polymerization
eukaryotes- about 14, pol alpha places primers and dissociates, pol E works on leading strand and pol S on the lagging strand
What other enzymes and accessory proteins aid in DNA replication
helicase, DNA gyrase, SSBPs, primase, DNA polymerase I, DNA ligase
List the 11 things to know about DNA replication, Draw a map of the process
- bidirectional
- helicase-separates the strands
- topoisomerase- reduces the torsional strain
- SSB proteins stabilize the strands
- RNA primer is needed to start
- DNA polymerase III adds nucleotides in the 5’–>3’ direction
- semi-discontinuous
- DNA polymerase removes RNA primer and replaces it with DNA
- DNA ligase seals the nick, after primer is replaced
- DNA polymerase I and III have 3’-5’ proofreading exonuclease activity
- DNA polymerase has 5’-3’ repair
what do each of the other enzymes and proteins do in DNA replication
helicase- separates DNA strand
DNA gyrase-relaxes supercoiling, it is ahead of the fork
SSBPs- single stranded binding proteins bind to single stranded DNA, the stabilize the isolated strands
primase-synthesizes the primers
DNA polymerase-removes primers, synthesizes DNA that replaces it in the 5’-3’ direction
DNA ligase-joins the DNA fragments with phosphodiester bonds
what do nucleases do
break phosphodiester bonds based on substrate
DNase
RNase
how are nucleases categorized
by bond location
endonuclease- inside
exonuclease-on 5’ end or 3’ end
what are telomers and what do they contain
unique, noncoding, repeated DNA sequences
why are telomers needed
because without telomers the DNA strand would shorten every time it is replicated.
Explain how DNA polymerase finishes replication
removes the primers
adds to the missing gaps
the 5’ end of the lagging strand has a gap that is never filled
the leading strand is longer-called 3’ overhang
what happens if telomers get too short
the repeated sequences can be added back by enzyme telomerase
how is a telomere added to the end of the DNA strand
*telomerase has RNA sequence in it
-the 3’ overhang is lengthened, telomerase binds to the longer (leading) strand and extends it
-telomerase is translocated, leaves
-the complement to the overhang is synthesized:
a primer is placed by primase complementary to the longer strand
-DNA polymerase and ligase fill in the gap
- a small gap is left at the end of the telomere
-proteins associate with the telomere to form a protective cap
how much % of DNA ends up being expressed?
2%
define what a gene is
sequence of DNA that codes for a functional product (like a polypeptide)
Describe transcription and translation, what are the final products
transcription: a sequence of nucleotides in a gene is used as a template for synthesis of RNA. the product is messenger RNA
Translation: a sequence of nucleotides in an mRNA strand directs the synthesis of a polypeptide.
What are the differences between DNA and RNA
- OH on pentose in RNA, H in DNA
- RNA has U instead of T
- RNA is usually single stranded, it can easily H bond to things. The secondary structure is a hairpin.
Define rRNA, tRNA snRNA and miRNA
rRNA-ribosomal, make up part of the ribosome
tRNA- transfer between mRNA and AA’s during translation
snRNA- small nuclear RNA, has function in pre-mRNA splicing
miRNA-microRNA, involved in regulation of expression
Explain how mRNA and tRNA can be specific
mRNA has a triplet codon that calls for a specific AA
tRNA has a nucleotide sequence within it (anticodon), another region covalently bonds to the corresponding AA
What are the components of ribosomes in prokaryotes and eukaryotes
prokaryotes (bacteria) monosome 70S, large (50S) and small (30S) subunit
eukaryotes: monosome 80S large (60S) and small (40S) subunit, overall has more proteins in subunits,
Both made of ribosomal proteins.
how is the transcription start site and the downstream genes labelled? what direction does it go in?
+1 start site
+2, +3.. in the direction of transcription
How are the upstream genes labelled
-1-2-3, in the direction opposite of transcription
What are the steps of Transcription in bacteria
initiation, elongation an termination
Describe the holoenzyme complex
RNA polymerase core enzyme with a sigma factor attached. it is the active complex that allows transcription to begin
what do B, B’ and sigma do in a holoenzyme
B and B’ porvide catalytic basis and the active site for transcription
sigma- is regulatory in the initiation of transcription.
Describe the parts of RNA polymerase in bacteria
made of alpha, beta, beta’ and w sub units. The core enzyme include all of these.
overall it transcribed DNA
Describe initiation in bacteria
*no primer is used RNA polymerase can just start
the promoter (upstream of the transcription site) is recognized by RNA polymerase core enzyme binds to the promoter
and the sigma factor of RNA polymerase binds to the consensus sequences separating the DNA strand around -10
after a few nucleotides, the sigma factor dissociates and the core enzyme continues to elongate the chain
what are the consensus sequences
homologous in different genes of the same organism in bacteria they are -35(TTGACA for transcription rate) and -10 (TATAAT/pribnow box for initiation)
Describe elongation in bacteria
RNA polymerase moves along template strand 3’->5’ making a temporary DNA/RNA duplex
the complementary RNA chain is polymerized from 5’->3’
how is the initiation of transcription specific?
there are alternative versions of the sigma factor that recognize different promoter sequences.
Describe termination in bacteria
RNA polymerase encounters the terminator sequence, a transcript (tail) is formed on RNA pol
2 possible types of termination
intrinsic: RNA pol tail has a GC region making it fold back on itself, creating hairpin.
hairpin cause RNA pol to stall, the end of the template strand has AT sequence, weak bonds that break once RNA pol with its tail tries to move past it, allowing the RNA polymerase to dissociate
rho dependent termination: termination factor rho that dissociates RNA hairpins and RNA-DNA interactions. Ro binds and chases the RNA polymerase. at the hairpin terminator sequence RNA pol stops and rho catches up, dissociates RNA pol and breaks the bonds between the template and the transcript.
what is different about bacterial mRNA?
polycistronic
What are the 4 major differences of transcription in eukaryotes
- happens in nucleus and mRNA are transported to cytoplasm to be translated
- Initiation requires remodeling of new chromatin
- initiation is more complex and requires protein factors
- The products of transcription have to be processed to produce mature mRNA
What are the RNA pols in eukaryotes and which one is most important
Pol 1, 2 and 3
pol 2 transcribes mRNA
Describe transcription in eukaryotes
Initiation:
TATA box(-25) and transcription factors GTFs and TFs are needed.
TFIID binds to TATA box sequence, other transcription factors and RNApol2 bind forming a pre initiation complex.
Termination: a sequence signals the RNApol2 to phosphorylate and dissociate.
Describe the spliceosome mechanism
- takes care of larger introns.
- made of snRNAs (small nuclear) rich in U residues. snRNAs form complex with proteins called snRNPs (small nuclear ribonucleoproteins)
- specific sequences (donor and acceptor) and consensus sequences attract specific snRNAs of the spliceosome. once the complex becomes snRNPs splicing starts.
- 3’ A residue attacks the 5’ splice site, cutting the chain. intermediate snRNP structure forms and links the cut 5’ end to the A. this forms “lariat” which can carry introns away.
- exons are ligated and the snRNPs release
What is the purpose of having introns and exons
alternative splicing can allow for different mRNA sequences to be formed from the same premRNA
which mRNA strand is identical to the coding strand?
the non template strand
Describe mRNA processing in eukaryotes
- M7G cap is added to 5’ end of mRNA, protects it from nuclease attack.
- polyA tail added to 3’ end also protects from nuclease
- introns are removed , by process of splicing by the spliceosome.
exons joined by ligase. Some introns (ribozymes) can take themselves out without enzymes.
the result is mature mRNA
Describe the one-gene enzyme hypothesis
experiments with bread mold where mutations were induced by radiation, the mutation was identified to be caused by a missing enzyme, proving one gene specifies one enzyme.
what are codons?
nucleotides in an mRNA that are decoded in groups of 3
how many possible reading frames are there in mRNA, why does it matter?
3 possible frames, if the frame is incorrect the translation will fail to produce the correct protein.
Explain the Crick and Brenner mRNA experiment
they experimented on bacteriophages that affect the growth rate of the bacteria they infected. They found that mutations of 3 sequences produced a nearly identical phenotype. demonstrating the existence of 3 bp codons.
what are the characteristics of the genetic code
nonoverlapping-ribonucleotides are part of one codon, no overlap.
non punctuating- there are no breaks between the codons
unambiguous-each codon specifies one AA
degenerate- an AA can come from more than one codon.
universal-codons specify the same AAs in all species with few exceptions.
what the 2 main features of transfer RNAs
- the 3’OH end is the acceptor end where the AA binds
2. the 5’ end is the anticodon loop that is complementary to an mRNA codon.
Describe the tRNA + AA rxn
aminoaclyation attaches an AA to the 3’ end of the tRNA. they attach at the carboxyl group of the AA. the charged tRNA can now start translation.
the chart given with the AAs that are produced by the nucleotide sequences are of ?
mRNA codons NOT tRNA anticodons
what are wobble base pairs?
they can form between the 3’ end of a codon and the 5’ end of an anticodon, where the bps are not AT GC
What are the 4 important features of the interior of the ribosome in eukaryotes
- There is a tunnel where mRNA can thread through
- There are 3 binding sites
E site exit: deacyl tRNA (nothing attached to acceptor end)
P site peptidyl tRNA
A site aminoacyl tRNA - There are 2 functional centers
decoding center
peptidyl transferase center - It has a polyprotein exit tunnel.
what is the only tRNA that can bind to the P site of the ribosome
tRNAmet binds to AUG in p site
describe translation initiation in bacteria
- IFs 1,2 and 3 bind to small subunit of ribosome to attract mRNA
- fMet-tRNA binds to AUG sequence of mature mRNA in the psite of the ribosome.
- shine-dalgarno sequence base pairs with 16SrRNA
- IF2 sets the reading frame
- IF3 leaves and the large subunit attaches to the complex. IF1 and 2 are released
Describe translation elongation cycle in bacteria
each cycle adds 1 AA.
- aminoacetyl- tRNA carries the next AA to A site. the charged tRNAs are carried by EF-Tu, hydrolysis releases bound tRNA
- a peptide bond forms between the 2 AAs in the P site
- EF-G helps move the peptidyl tRNA from A-P site an the deacyl-tRNA from P-E site, they dissociate
- a third charged tRNA is ready to enter the A site. pp chain grows in P site
Describe translation termination in bacteria
- the stop codon is reached, RF1 or 2 binds to the stop codon in the A site instead of a tRNA
- ribosome subunits separate and mRNA, pp chain and RF factors release. pp folds into conformation, tRNA releases
what is the preinitiation complex in eukaryotic translation initiation called? the initiation factors?
43-S initiation complex
eIFs
what is the mRNA complex in translation initiation of eukaryotes made of and what does it do
eIF4G
eIF4E
polyA binding protein
brings 5’ end of mRNA around and creates circular path for translation
describe translation in eukaryotes
Initiation: tRNA and ribosomes move in 5’-3’ direction and scan the 5’ UTR for the AUG sequence
eIFG binds M7G cap to 5’ end of mRNA
polyAA tail added by eIFG.
Elongation: translation occurs along the loop
Termination: the stop codon is reached, ribosome might be recycled for use again.
What direction does AA translation go in
N–> C
how are R groups categorized
non polar-hydrophobic (interior of protein)
polar-hydrophillic (exterior of protein)
polar charged: + or - charge, also on surface of proteins
name 2 RNA enzymes that are not proteins
spliceosome
ribosome
Describe protein structure
primary-AAs
secondary- alpha helix and beta pleated sheet
3ary-3D structure
4ary-several 3D structures together
what bonds stabalize each protein structure
2ndary- H bonds between bps on backbone
3ary- H bonds between R groups
at what structure do proteins become functional
3ary
at what structure do proteins become functional
3ary
name an example of 4ary proteins
hemoglobin
what causes sickle cell anemia
AA substitution of Val instead of Glu acid. exposes the hydrophobic region of Hb and RBCs get sickle cell shape, they also stick to each other. all caused by a gene mutation
what is constitutive gene expression
when genes are expressed in all conditions, housekeeping genes encode proteins needed for survival
what proteins might housekeeping genes encode
ribosomes, enzymes, rRNA’s mitochondrial enzymes, histone proteins, tRNA
why is expression of genes regulated
allows the cell to produce the proper proteins, at the proper time in proper amounts
inducible gene regulation
expression turned on
goes from off->on
repressible gene regulation
expression turned off
goes from on->off
in bacterial gene regulation what proteins control gene expression
repressors and activators
when does bacterial gene regulation occur
mostly during transcription
what do repressors do
bind to operator DNA sequences and prevent transcription (negative regulation)
what do activators do
bind to activating binding site and stimulate transcription (positive regulation)
when can inducible translation be +? -?
- repressor could be bound blocking transcription
inducer binds to repressor, stops repressor, transcription continues.
+ inducer binds to inactive activator, activating transcription
when can repressors be + and -?
- corepressor binds to inactive repressor, allowing it to stop translation
+ corepressor binds to activator, inactivating it and stopping transcription
inducible vs repressible
negative vs positive
inducible (off->on)
repressible (on->off)
negative (repressors-prevents)
positive (activators-stimulation)
what are the 3 genes involved in lactose metabolism
lacZ, lacY and lacA
if lactose is not available, the operon is ______
repressed
if lactose is available the operon is _____
induced
what are the products of each gene in lac operon
Z-galactose
y-lactose permease
A-galactoside
what is the role of allolactose
when lactose is available, it is created and binds to the repressor that stops lactose metabolism
when lactose is not available, allolactose levels decrease and they are released from repressor, lactose metabolism becomes repressed
If glucose levels are low, the lac operon is ???
maximally expressed
if glucose levels are high the lac operon is ____
not stimulated even under inducible conditions
what does the CAP-cAMP complex do?
together they are activating proteins they stimulate RNA polymerase to bind, to transcribe lacmRNA
What are the 3 types of eukaryotic gene regulation?
- regulation of chromatin structure
- regulation of transcription
- post transcriptional regulation
what are the 4 ways chromatin structure is regulated
- DNA methylation
- histone modification
- nucleosome composition
- nucleosome remodelling
how are histones modified
the tails are modified, AA in tails are acetylated, methylated, phosphorylated to open the chromatin
what is DNA methylation
the removal of methyl groups from the N bases in DNA. represses gene expression
explain what the difference between open and closed chromatin is
open- allows for transcription
closed-transcription and any modification is stopped
explain what the difference between open and closed chromatin is
open- allows for transcription
closed-transcription and any modification is stopped
explain how nucleosome composition regulates gene expression
histone variants are placed in the nucleosome changing the positioning of them
what is nucleosome remodeling
repositioning or removing nucleosomes
what are the 2 types of transcription regulators (euk)
activators and repressors
what are the main characteristics of transcription regulators (euk)
- over large distances
- position and orientation independent
- bind to enhancers/silencers and affect small subsets of genes
how do activators regulate transcription in euk
they bind to enhancers to recruit and position GTFs and RNAPII at a promoter
how do repressors regulation euk transcription
bind to silencers and decrease transcription initiation
what are the 3 ways repressors affect transcription in euk
- block the binding of an activator
- inhibit the formation of preinitiation complex
- recruit or stimulate chromatin remodeling that leads to closed chromatin
what are siRNA’s
small interfering double stranded RNA in cytoplasm
Explain what RISC is
RNA induced gene silencing, a type of post transcriptional control where DICER takes over the complex, leaves RNA single stranded, attracts other single strand to bind and both end up degraded
