Exam Review Flashcards
1
Q
Watson and Crick suggested that the double helical DNA model suggests a _________ copying mechanism which means…
A
Semi conservative
after one round of DNA replication, each DNA contains 1 parental and 1 newly syntheized strand
2
Q
What are the 3 models of DNA copying mechanisms
A
- Conservation replication
- Dispersive Replication
- Semi-conservative Replication
3
Q
What are the 3 modes of DNA replication
A
- Theta replication
- Rolling circle replication
- Linear chromosome replication
4
Q
Describe theta replication
A
- Replication that occurs in mostly circular DNA
- double stranded DNA unwinds at the replication origin
- This produces single stranded templates DNA synthesis and replication bubble forms with a replication fork at each end
- the fork then proceeds around the circle bidirectionally
- Eventually two circular DNA molecules are produced
5
Q
Rolling Circle Replication
A
- Specialized form of replication that occurs in the F factor and some viruses only
- replication is initiated by a break in one of the nucleotide strands
- synthesis begins at the 3’ end of the broken strand; the inner strand is then used as a template. the 5’ en d of the broken strand is displaced
- cleavage releases a single stranded linear DNA and a double-stranded circular DNA. Replication is unidirectional.
- the linear DNA may circularize and sere as a template for the synthesis of a complementary strand
6
Q
linear replication
A
- Linear chromosome replication occurs in the linear chromosomes of eukaryotic cells.
- This DNA replication is bidirectional.
7
Q
What are the requirements for DNA replication (4)
A
- DNA dependant DNA polymerase
- Four deoxyribonucleoside triphosphates (dNTPs)
- A single stranded template of DNA to be copied
- An RNA primer (provides the 3’ OH end to initiate DNA synthesis by DNA polymerase)
8
Q
DNA is ALWAYS synthesized in what direction?
A
5’ -> 3’
9
Q
DNA synthesis is ____ on the leading strand and ___ in the lagging strand
A
continuous, discontinuous
10
Q
Fragments formed during synthesis of the lagging strand is called
A
Okazaki fragments
11
Q
What are DnaA proteins involved in prokaryotic DNA replication
A
- They are initiator proteins that bind to oriC (the origin)
- This causes a short stretch of DNA to unwind
12
Q
DNA Helicase (Prokaryotes)
A
- Unwinds the DNA in the 5’ to the 3’ direction
- Only travels on the lagging strand
13
Q
Single Strand binding proteins (Prok.)
A
Coats unwound single stranded DNA to protect and keep DNA single-stranded (prevents hairpins)
14
Q
DNA Gyrase (prok.)
A
- A topoisomerase that makes double stranded breaks in DNA to relive the torsional strain that builds up from the activity of the DNA helicase.
15
Q
____ requires a primer to initiate synthesis ___ does not
A
DNA polymerization, RNA polymerase
16
Q
Primase (RNA Polymerase) (prok.)
A
Synthesizes short RNA primer that provides the 3’ OH end for DNA polymerase to begin DNA synthesis
17
Q
Primase binds to ___ to initiate synthesis
A
Helicase
18
Q
DNA Polymerase (prok.)
A
Elongates the new polynucleotide strand by catalyzing DNA polymerisation.
19
Q
T or F: DNA polymerase requires a primer
A
True
20
Q
DNA Pol III (prok.)
A
- The main DNA Pol
- has 5’-> 3’ polymerase activity and 3’ to 5’ exonuclease activity.
- can add dNTP on the 3’ end and also back up to remove a nucleotide that has been mis incorporated
21
Q
DNA Pol I (prok.)
A
- Similar to DNA Pol III, but has 5’ -> 3’ exonuclease activity so it can remove the RNA primer and replace it with DNA
22
Q
DNA Ligase (prok.)
A
Makes phosphodiester bonds between 5’ phosphate and the 3’ OH froup and seals the nicks in the lagging strand
23
Q
Helicase (Euk.)
A
- Binds to initiator protein on double streanded DNA
- involves the MCM complex of proteins
24
Q
In eukaryotes DNA replication only occurs during ___ phase
A
S
25
Polymerase Alpha (euk)
- Has primase activity
- generates the RNA primer
26
Polymerase Epsilon (euk)
preforms leading strand replicaiton
27
Polymerase delta (euk)
Preforms lagging strand replicaition
28
when DNA is doubled in euk. histones must be ___ and the modifications on them need to be replicated
doubled
29
The telomere problem
- The chromosome end will be degraded causeing shortening every round of replication.
- leaves gaps in DNA that are not replicated causing shortening
30
Telomerase
- Activity extends eukaryotic chromosome ends in replicating cells
- can extent the 3' end without use of complementary DNA template
- results in shortening telomeres over time
31
Shorter telomeres is associated with...
Cellular senescence and death
32
What makes RNA less stable than DNA?
The presence of 2'OH groups in ribose causes it to react intramolecularly with the 3'OH site resulting in phosphate breakage.
33
What is transcription
Synthesis of RNA from DNA
34
In what Direction is RNA synthesis
RNA is synthesized in the 5' to 3' direction using the 3' to 5' DNA template strand.
35
The precursors to RNA syntheis are..
Ribonucleoside triphosphates (rNTPs)
36
RNA Polymerase Holoenzyme Subunits
- alpha
- beta
- beta-prime
- omega
37
What is the function of the alpha subunit?
Involved in the assembly of the tetrametric core
38
what is the function of the beta subunit?
Contains the ribonucleoside triphosphate (rNTP) binding site
39
What is the function of the beta prime subunit?
Contains the DNA binding region
40
What is the function of the omega subunit?
It helps to stabilize the tetrameric core
41
What is the function of the sigma subunit
- it binds to the RNA polymerase tetrametric core and assists in the correct initiation of transcription specifically at the promotor region of the prokaryotic gene. Many types of sigma factors -> allow for specificity.
- recognizes and binds to the -35 and -10 consensus sequances in the promotor region, properly positioning the RNA polymerase to begin transcription
42
Why is the -10 consensus sequence prone to unwinding?
Due to its AT rich content
43
Describe initiation of transcription in prokaryotes
1. holoenzyme binds to the promoter and RNA polymerase is positioned over the transcription start site and has unwound the DNA to produce a single stranded template
2. the orientation and spacing of the consensus sequence on a DNA strand determine which strand will be the template for transcription and therby determine the direction of transcription
3. RNA polymerase binds, unwinds and joins first 2 nucleotides
44
Does RNA synthesis require a primer
no
45
Elongation
1. Occurs when sigma factor is released and RNA polymerase begins to move along the 3' to 5' DNA template strand.
2. Complementary nucleotides are added
3. Localized DNA unwinding ahead of RNA polymerase and generates a "transcription bubble"
4. Transcription bubble moves along with the unwound DNA rewinds behind the bubble
46
RNA polymerase has both helix ___ and ___ activities
unwinding, rewinding
47
Termination
- stop when RNA polymerase reaches the terminator region of the gene
- occurs upstream of where the actual termination takes place
- the newly synthesized RNA together with the RNA polymerase are released
48
What are the two major types of terminators in bacterial cells?
Rho-dependant (requires the Rho factor) and Rho-independant (aka intrinsic terminators)
49
Describe Rho-dependant Termination
Sequence Features:
1. DNA sequence of terminator site causes polymerase to pause
2. DNA sequance upstream of terminator encodes a stretch of RNA that is C rich and devoid of secondary structure
a) called the rho utilization
b) rho binds to rut site
- Rho moves alonf RNA towards paused polymerase
- rho factor has helicase activity
- unwinds the RNA-DNA hybrid
- ends transcription
50
Describe Rho-independent termination
Common features
1. contains inverted repeats
2. a string of 6-9 A's follows the inverted repeats
- poly A sequance is transcribed into a poly U tail after hairpin is transcribed
- causes polymerase pause
- hairpin forms and destabilizes the DNA-RNA hybrid
- assisted by weak A-U base pairing
51
true or false: Pol I, II, and III are inly recruited to their promoter specific accessory proteins
true
52
Initiation in Eukaryotes
Involves a step wise assembly of general Transcription Factors of Pol II
- Tata box is the fist to assemble then followed by the remaining general TFs and Pol II
- assembly of the TFs and Pol II causes 11-15 bp of surrounding DNA to unwind
- template strand positioned within active site of RNA pol
53
Is the pre-initiation complex sufficient to initiate transcription?
yes
54
what is a mediator
involves multi-subunit that permits interactions with other activator/repressor proteins bound to upstream/downstream regulatory regions or enhancer sequences
55
Elongation in Eukaryotes
- Polymerase moves along the template strand leaving transcription factor intact at the promoter for reinitiation of transcrption with new polymerase
- RNA pol maintains 8nt transcption bubble
- DNA-RNA hybrid bends at right angle
- positions the OH group at the active site where nucleotides are added to the 3' end of the growing chain
- newly sythesized RNA is seperated from DNA and exits through another cleft
56
Termination in RNA Pol I
Requires a termination factor similar to the rho factor in prokaryotes
57
Termination in RNA Pol III
Ends after transcribing a terminator sequence that produces a string of Us that is downstream of hairpin. Similar to rho-independant termination
58
Termination in Pol II
Transcription continues past termination sequence
- RNA is cleaved at consensus sequence in the RNA
59
Termination in Eukaryotes
- Cleaved RNA results in 2 RNAs: one that will encode a protein and the other with its 5' trailing out of the RNA polymerase
- Rat 1 (5' -> 3' exonuclease) attaches to the 5' end and chews up the RNA
- rat 1 does not bind to the 5' end of the preotein coding mRNA because it is protected by a 5' cap
60
What is Colinearity
- the coding region in prokaryotes that is uninterupted
- The sequance of the gene directly corresponds to the amino acid sequance
the number of nucleotides is proportional to the number of amino acids (3:1)
61
The shine delgarno sequance is found in ___ (prok or euk)
Prokaryotes
62
What are the three primary regions of a mature mRNA
5' Untranslated Region (5'UTR): Does not code for amino acids, binds the ribosomal complex
Protein Coding Region: Comprises the codons that specify the amino acids begins with a start codon and ends with a stop codon
3' UTR: does not code for amino acids. Affects the stability of the mRNA and regulates its translation
63
Exons are...
Protein coding segments
64
Introns are...
inervating (non-coding) segments
65
removal of ___ along with additional RNA processing steps are required to form the mRNA that will be translation into ___
introns, polypeptide
66
What is required to remove the intron?
the splicosomal complex which contains small nuclear RNAs and proteins
67
Where does splicing take place?
In the nucleus
68
What are the 3 main processing steps in eukaryotic nuclear pre-mRNA
1. the addition of 7-methyl guanosine cap. linked betwen the 5' phosphate of the 7 MG and the 5' phophphate of the first RNA
2. addition of a poly A tail to the 3' end by poly A polymerase
3. Removal of introns from pre-mRNA called splicing
69
Describe the 5' cap
Methyl groups are added to:
- 7' position of the terminal guanine
- 2' OH group of the sugar of the first and sometimes second nucleotide
- to adenine if it is the second nucleotide
70
Descibe the 3' poly A tail
50-250 As added to the 3' end
- A are added without a template (polyadenylation)
- Poly A consensus sequance (AAUAAA) is the polyadenylation signal located 11 to 30 nucleotides upstream of the cleavage site
- affects stability of mRNA, facilitates attachment of the ribosome and export out of the nucleous
71
What is the polyadenylation signal
AAUAAA
72
Descibe RNA splicing
- removal of introns from pre-RNA
- Removal of introns must be precise in order to properly fuse the 3' end of one exon to the 5' end of the next exon
73
How does the eukarotic cell ensure that introns are properly removed?
Every intron has 3 conserved sequances thata re required for its precise removal:
1. 5' splice sequances containing the junction seequances GU (5' end of the intron)
2. 3' splice sequence containing the junction sequence AG
3. Intron branch point: a conserved A residue is located 18-40 nt upstream of 3' splice site
74
What are the Steps of RNA splicing
1. Pre-mRNA is cut at the 5' splice site and the 5' end of this intron attaches to the branch point (Lariat)
- 5' phosphate of the G nucleotide binds with the 2' OH group of the A nucleotide at the branch point
2. Cut is made at the 3' splice site and simultaneously the 3' end of exon 1 becomes covalently attached to the 5' end of exon 2
- intron is released as a lariat and degraded
75
Describe a Spliceosome
- RNA/ pritein structure
- contains 5 small nuclear RNAs
- The snRNAs associated with about 300 small proteins to form small nuclear SNURPS
76
What is alternative splicing?
- an alternative processing pathway
- pre-mRNA is processed in different ways to produce alternative types of mRNA
- alternative exons are used
- results in different proteins from the same DNA sequance (unless in the UTR region)
77
Describe the concept of multiple 3' cleavage sites?
- alternative procssing pathway
-Can generate longer or shorter exons
- may or may not produce a different protein. depends if upstream or downstream of stop codon
- note that this is not the same as the 3' splice site
78
What is Calcitonin
an alternative processing pathway
regulates calcium concentration in the blood
79
what is CGRP
an alternative processing pathway
Causes dialation of blood vessels and the transmission of pain
80
How is RNA editing carried out?
- Carried out by guide RNAs that can be made in the cell
- gRNAs direct the insertion of urodine bases into the mRNA by a repair polymerase
- This perminatly modifies the mRNA by making new codons that specify new amino acids
81
What is the editing of Apoplipoproteins B mRNA
- Apoplipoproteins are blood proteins that carry lipids
- RNA editing by cytidine deaminase changes C to U converting a normal glutamine codon to a termination codon which truncates the protein and gives it a different function with respect to lipid binding
82
Describe tRNA modifications
- the anticodon of the tRNA base pairs with the codon of mRNA
- The amino acid is covalently attached to the 3' end of the tRNA
- tRNAs often contain modified ribonucleotides through the action of tRNA modifying enzymes
83
What processing events do tRNAs undergo
- removal of extra 5' and 3' sequances
- removal of introns
- nucleotides 5'-CCA-3' are added to the 3' end of the tRNA for all tRNAs
- modification of several nucleotides
84
What is a ribosome
- comprised of 2 subunits and has a key role in protein synthesis including the formation of peptide bonds between amino acids
85
Where does rRNA sytheisis occur in both prok. and euk.
Prokaryotes
- No nuclear envelope so rRNA synthesis and ribosome assembly occur in the cytoplasm
Eukaryotes
- nucleus is the site of rRNA synthesis and ribosome assembly
86
snRNAs
Splices introns from genetic transcripts
87
snoRNAs
- act in complezes with proteins
- in euk., guide the enzymatic chemical modifications of ribosomal RNAs, transfer RNAs and small nuclear RNAs
88
siRNA, miRNA
- found in euk.
- act as short, single stranded RNAs that bind to complementary sequances in mRNA
89
Crispr RNA
- found in prok.
- encoded by DNA sequances found in prokaryotic genomes
works in association with the prokaryotic cas 9 nuclease to cleave foreign DNA that might happen to enter a host cell
90
LncRNA
- known to function in eukaypotic cells
- Longer than miRNAs
- regulate and control gene expression at the level transcription or translation by binding mRNA or sequestering micro-RNAs that control gene expression
- bind or recruit proteins involved in DNA modification
91
What is sickle cell anemia?
It is caused by single acid change in one of the protein chains that make up hemoglobin red blood cells
92
true or false: transcription and translation often occur simultaneously in prokaryotes
true
93
where do transcription and translation occur in eukaryotes?
the nucleolus and cytoplasm respectively
94
What is the one gene, one colinear polypeptide theory
The sequance of base pair triplets in the coding region of a gene specify a colinear sequance of amino acids in its polypeptide product
95
Structure of Amino acids
- Proteins are made of polypeptides
- amino acids has a free amino group (N-term), a free carboxyl group (C-term), and side group R
96
How are amino acids joined
they are joined covalently by peptide bonds
97
Why is the genetic code a triplet code?
4^3 cause that is the only way there was enough codons to encode for 20 amino acids
4^3 = 64 therefore there is degeneracy in the code
98
How was the genetic code determined
1. Synthetic mRNAs of repeated sequance (homopolymers) were tested in vitro for protein synthesis: produced homopolypeptides
2. Each individual amino acid was tested until the polypeptide set off the geiger counter
3. this confirmed UUU specifies phenylalinine
4. they then moved on to the next homopolymer and repeat the process
99
What is the wobble position
Flexibility in the binding at the 3rd codon position (1st anticodon.
- This permits the tRNA anticodon to bind codons that bind the same amino acid even if the sequance isnt complementary to the tRNA
100
What feature in the genetic code explains degeneracy
Oftentimes the 3rd codon position can be changed and still specify the same amino acid
101
What are the 4 Macromolecules of Translation
1. Ribosomes/ ribosomal RNA
2. Amino acid activating enzymes
3. tRNA molecules
4. soluble proteins involved in polypeptide chain initiation, elongation and termination
102
What is a Ribosome?
- Ribosomes are composed of proteins and several diff rRNAs
- Ribosomes are composed of both a large and small subunit
- a ribosome is an RNA machine with key roles in protein synthesis including the formtion of peptide bonds between amino acids
103
What is Aminoacyl tRNA syntheses?
- this enzyme charges the tRNA with its specific amino acid
- an amino acid becomes attached to the appropriate tRNA by the amino-acyl tRNA synthetade in a two step rxn:
1. amino acid reacts with ATP
2. the amino acid is tranfered to tRNA and AMP is released
104
What are tRNAs
- tRNA are adapters between amino acids and the codons in mRNA
- the anticodon of the tRNA base pair with the codon of mRNA
- The amino acid is covalently attached to the 3' end of the tRNA
105
Translation initiation (prok)
1. 16S of the 30S small ribosomal sunbunit contains the complement to the shine-delgarno sequance in the mRNA. Pairing of the two sequances positions the ribosome near the AUG start codon
2. IF-3 inhibits the large subunit from binding (30S) small subunit
3. IF-1 and IF-2 position fMet-tRNA over the start codon
4. IF-3 dissociated allowing the 50S subunit to bind
5. GTP is hydorlyzed to GDP and the remaining IF factors dissociate
6. complete comllex is now called 70S initation complex
106
What. is the difference between prok. and euk. translation initiation
1. fmet in prok. just met in euk.
2. no shine delgarno sequance in euk.
3. Kozak sequance is used in place of the shine delgarno sequance
4. poly A tail of the euk mRNA interacts with the mRNA 5' 7-MG cap structure via the cap binding protein complex to promote translation initiation
107
Translation elongation
1. A charged tRNA binds to the A-site of the ribosome
2. once in the A site, GTP is hydrolyzed to GDP and is released along with EF-Tu
3. rRNA in large subunit catalyzes formation of a peptide bond
4. chanin is transfered to the aa in the A-site
5. rRNA subunit catalyzes formation of the peptide bond
6. releases the tRNA in the P site from its AA
108
what site does the F-Met or first Met in a protein strat in during protein synthesis
the P-site
109
what direction does the ribosome move does the mRNA
5' -> 3'
110
What direction does the polypeptide chain grow
N-term -> C-term
111
Translation Termination
Termination occurs when stop codon enters the A-site of the ribosome?
1. when stop codon is encountered a release factor (RF) binds to the A-site
2. RF-1 or 2 binding alters activity of the peptidyl transferase, releasing the polypeptide and leading to the termination.
3. the binding of RF 3 and GTP to the ribosome assists in the dismantling of the entire complex
112
What stop codons do RF-1 recognize
UAG, UAA
113
what stop codons does RF 2 recognize
UAA, and UGA
114
what does Chloromycetin inhibit?
Formation of peptide bonds
115
what does Erythromycin inhibit
translocation of mRNA along the ribosome
116
what does Neomyacin inhibit
Interactions between tRNA and mRNA
117
What does Streptomycin inhibit
initiation of translation
118
what does tetracycline inhibit
binding of tRNA to ribosome
119
what does Paromomycin inhibit
Validation of mRNA-tRNA match
120
Define a Mutant
An organism that carries one or more mutations in its genetic material
121
True or False: Mutations are the source of all genetic variation
true
122
During meiosis, recombination between __ chromosomes rearranges genetic variability into a new combination
homologous
123
Somatic Mutations
Occur in somatic cells; it will occur only in the descendants of that cell and will not be transmitted to the progeny.
124
Germinal Mutations
Occur in germ-line cells and will be transmitted through the gametes to the progeny
125
What are the 4 main types of Gene Mutations?
1. Base Substitutions
2. Insertions and Deletions
3. Tautomeric Shift
4. Expanding nucleotide repeats
126
Describe a transition Mutation
Replaces a pyrimidine with another pyrimidine, or purine with purine.
127
Describe a transversion mutation
replaces a pyrimidine with a purine or vise versa
128
Describe insertion and deletion mutations
Add one or two base pairs that alter the reading frame of the gene distal to the site of the mutation.
129
What is a tautomeric shoft mutation?
The movement of H atoms from one position in a purine or pyrimidine base to another.
- This is a rare mutation that can occur spontaneously during DNA replication where they alter DNA base pairing
130
Can tautomeric shifts cause spontaneous mutations?
Yes
131
Can a tautomeric shift generate A:C and G:T pairs?
yes
132
Describe expanding nucleotide repeats
- Expansion of triple repeats
- involves DNA replication
- Dynamic mutation because the repeat copy number is in flux with each round of replication.
- Mutations can be outside of the coding region that affects its expression
133
Forward Mutation
wildtype -> mutant type
134
Reverse Mutation
Mutant type -> wild type
135
Missense Mutation
amino acid -> different amino acid
136
Nonsense Mutation
Sense codon -> nonsense codon (stop)
137
Silent Mutation
codon -> synonoomous codon still specifying the same amino acid
138
Neutral Mutation
amino acid change with no observable change in protein function. The amino acid changes to one of similar chemical type
139
Loss of function mutation
Result of mutations that cause complete or partial loss of normal protein function
140
Gain of function mutation
is the result of a mutation that causes the cell to produce a protein or gene product whose function is not normally present
141
Conditional Mutation
Is expressed only under certain conditions
142
lethal mutation
causes pre mature cell death
143
Temperature sensitive allele
An allele whose functional product is conditional on the temperature
144
Supressor Mutation
A second site mutation that hides or suppresses the effect of the first mutation
145
Intragenic Supressor
Supressor mutations within the same gene
146
Intergenic suppressor
A suppressor mutation can be present within a different gene
147
Induced mutations
Mutations that occur as a result of exposure to external factors
148
Spontaneous mutations
Mutations that occur under normal circumstances as a result of internal factors
149
Spontaneous Replication Errors
- Tautomeric shifts
- Mispairing due to other structures
- Incorporated errors and replicated errors
- causes deletions and insertions
- spontaneous chemical changes
150
What causes deletions and insertions (2)
1. Strand slippage: can be insertion or deletion
2. Unequal crossing over during meiosis
151
Describe Depurination
- a spontaneous chemical change
- breakage of the covalent bond between the purine base and the 1' carbon atom of the deoxyribose sugar. creates an apurinic site
152
Describe Deamination
- spontaneous chemical change
- Deamination is a loss of amino group, typically from cytosine. This gives rise to uricil, which will pair with adenine during replication. In the next cycle it will pair with T. End result is C -> T
153
Mutagen
An environmental agent that increases the mutation rate above the spontaneous rate
154
Base Analogue (Mutagen)
Chemical with structures similar to any of the standard 4 mucleotides
155
What is 5-bromouracil an anolgue of
thyamine which causes G -> transition
156
what is 2-aminopurine an analogue of
Adenine which can mispair eith cytosine
157
What are Alkylating Agents (mutagens)
Mutagens that react with DNA bases and add methyl or ethyl groups
- EMA and mustard gas
158
Describe Deaminating Chemicals (mutagens)
- This is in addition to spontaneous deamination noted for spontaneous chemical changes
- Nitrous acid causes c->T and G->A transitions
159
Describe Hydroxylamine (mutagen)
- Vary specific base-modifying mutagen
- adds hydroxyl group to cytosine -> hydroxylaminocytosine
- increases occurence of rare tautomer that pairs with adenine
- only affects cytosine
- C -> T transition
160
Oxidative Radicles (mutagens)
- Reactive forms of oxygen
- producef through normal anerobic metabolism
- can also be produced by chemicals and radiation
- can affect different bases
161
Intercalating Agents (mutagens)
- sandwich themsekces (intercalate) between adjacent base pairs
- distorts DNA helix
- causes insetions and deletions -> frameshift mutations
162
Radiation (mutagen), 2 types of radiation
Ionizing radiation
- dislofge electrons from the atoms they encounter
- Stable atom -> freee radicles, reactive ions
- Breaks phosphodiester bonds leading to double stranded breaks
UV Radtion
- less intense than ionizing
- Pyrimidine bases absorb UV light
- induces chemical bonds between two adjacent pyrimidine molecules on the same strand of DNA
- thymidine dimers most frequant
163
What are the two general features of DNA Repair
1. Most DNA repair mechanisms require double stranded DNA: As one nucleotide is being repaired, the complementary strand is acting as a template for the repair
2. DNA repair has built in redundancy. Many types of DNA damage can be repaired by more than one repair system, helps ensure survival.
164
DNA Mismatch repair
- Old DNA has methyl groups attached to it, new DNA does not
- Allows the MMR machinery to differentiate the new versus original strand
- MMR degrades new strand between nick and mismatched bases
- Polymerases and DNA ligases fill the gap using the old strand as the template
- Similar processes between prok. and euk., but euk. dont have methylated DNA, so unknown how MMR recognizes old and new strands
- Mutations in MMR leads to elevated somatic mutations -> cancer
165
Direct DNA Repair
Retores the original structure:
1. photoreactivation of UV-induced pyrimidine dimers. Breaks the covalent bond in the dimers
2. Enzyme call O-methylguanine DNA methyltransferase removed the methyl grouo from the O methylguanine, restoring th ebase to guanine
166
Base Excision DNA Repair
- DNA glycosylases excises the damaged base or bases
- these enzymes are specific for each modified base
- just the base is removed not the deoxyribose sugar
- AP endonuclease cleaves the phosphodiester bond
- other enzymes remove the deoxyribise sugar
- A DNA polymerase fills in the gap using the undamaged complementaey strand of DNA as the template
- DNA ligase seals the brak left by DNA polymerase
167
Nucleotide-Excision DNA Repair
- NER is complex of enzymes that scans the DNA for distortions in the DNA
- DNA is seperated and stabilized by single stranded DNA Binding proteins
- Phosphodiester bonds on either side of mutation are cleaved
- damaged strand is removed by helicase
- gap is filled in with DNA polymerase
- ends are sealed by DNA ligase
168
Repair of Double stranded DNA breaks
- Caused by ionizing radiation, oxidative radicles and other DNA damaging agents
- stalls DNA replication and could lead to chromosomal rearrangements which in turn can cause deletions, duplications, inversions and translocations
Two paths repair these
1. Homology directed repair
2. nonhomologous end joing
169
Homology dependant double stranded DNA repair
- homologous recombination
- repairs a briken DNA molecule by using the genetic information contained in the sister chromatid
- Employs the same system as in homologous recombination that occurs during crossing over during meiosis
170
Non-homologous end joining in double stranded breaks
- Repairs double stranded breaks
- independant of homologous template
- proteins recognize broken ends of DNA and bind them together
- leads to deletions and insertions and translocations
- important in somatic recombination
- involves DNA polymerases, nucleases and ligases
171
The enzyme with a built-in RNA strand template, which enables DNA synthesis at the end of the eukaryotic chromosome, is:
Telomerase
172
What are transposable elements
Segments of DNA capable from moving from one location in a chromosome to another, or even to a different chromosome
173
General Characteristics of Transposable Elements
1. Create staggered breaks in target DNA
2. Transposable element attaches to single stranded ends of DNA
3. DNA is replicated at the gaps
- flanking direct repeats
- generated in the process of transposition
4. Terminal inverted repeats associated with the transposable elements
- inverted and complimentary
174
Terminal inverted repeats are recognized by .... that catalyze transposition
Transposases
175
There are ___ classes of transposable elements based on ___
2, DNA or RNA
176
Class I. Retrotransposons (RNA intermediate)
- DNA copy of element made by reverse transcription from its RNA and then inserted into a new chromosomal site
- requires a reverse transcriptase - reverse flow of genetic information from RNA to DNA
Two Types;
1. Retrovirus or Retrovirus like elements
2. Retroposons or retrotransposons
Both use replicative transposition
177
Class II. DNA transposons-catalyzed by transposase
1. Non-replicative transposons - element is physically cut out of one site in a chromosome or plasmid and pasted into a new site (cut and paste)
2. Replicative Transposons - element is replicated with one copy and inserted at a new site and one remains at the og. site (copy and paste)
178
How are transposable elements controlled
1. DNA is methylated where transposons are common
2. alterations in chromatin structure prevent transcription
3. Control of transposase translation by piwi-interacting RNAs (piRNAs). Bind to Piwi proteins and inhibit tranlation of transposase mRNA
179
what is the mutagenic effect (transposable elements)
- Transposition entails the exchange of DNA sequances and recombination
- Leads to rearrangements such as duplications, deletions, inversions, and translocations
180
mutanagenic effect can occur by pairing of direct repeat which leads to ___
Deletion
181
mutanagenic effect can occur by pairing of inverted repeat which leads to ___
Inversion
182
What are the transposable element groups in bacteria
1. Simple transposable elements
2. Complex transposable elements
183
Simple Transposable Elements
- only carry the information required for movement
- contain the transposase gene
- contain both terminal inverted repeats and direct repeats
184
Complex transposable elements
Composite transposons
- any sequence that is flanked by two insertion sequences
Noncomposite transposons
- lack insertion sequances
- posses the transposase gene
- have terminal inverted repeats
185
Ac-Activator Gene (transposons)
- Contains terminal inverted repeats
- has transposase gene
- autonomous transposition
186
Ds Dissociation gene (transposon)
- Similar to Ac
- Posses inactivated transposase gene
- Requires transposase from Ac to transpose
- Noautonomous transposition
187
Ac stimulates chromosome ___ at site of Ds
breakage
188
Transposable elements in Eukaryotes
p elements
- possess terminal inverted repeats
- DNA transposons
- contains both a transposase and repressor of transposition
- creates phenomenon of hybrid dysgenesis
189
What is Hybrid Dysgenesis?
- Occurs when p elements are introduced into a cell that does not have them in its genome
- sudden appearance of numerous mutations
- chromosome aberrations
- sterility
190
Describe the process of Hybrid Dysgenesis
- Repressor protein is a cytoplasmic protein that inhibits transposition
- inhibitor is in the egg of the female (P+)
- if this P+ is crossed with P- or P+ male, the inhibitor in the egg will prevent transposition and the fly will be normal or wild type
if a P- female is crossed with a P+ male the lack of inhibitor in the egg will alow for rapid transposition, leading to mutations and sterility
191
Sines - Transposable Elements
- not identicle
- have short flanking direct repeats when inserted into DNA
- arose through an RNA intermediate
- Alu is a short interpersed elements
192
Lines - Transposable Elements
Long interspersed elements
193
Constitutively expressed Genes
- Essential components of almost all living cells
- Genes that specify products of this type are continuously expressed in most cells
194
Inducible and Repressible Genes
- Some gene products are needed for cell growth only under certain environmental conditions
- Regulatory mechanisms allow the synthesis of these gene products only when they are needed (inducible) or shut down genes when they are no longer need (repressable)
195
Transcription in bacteria is regulated by ___
operons
196
Negative regulatory proteins ___ transcription
Positive regulator proteins ___ transcription
inhibit, activate
197
Inducible operon
When transcription of the operon is normally off and something happens that turns it on.
198
Repressible Operon
Normally turned on, and something happens that it needs to be turned off
199
In a negative inducible operon:
Regulator protein is a ___ and blocks the binding if the RNA polymerase to the ___
inhibitor, promoter
200
Negative Inducible Operon
- Transcription is normally turned off and must be turned on
- Repressor needs to be relived of its duties in order for transcription to proceed
- inducer activates it
- repressor can no longer bind to DNA
Negative: Regulator protein is an inhibitor or repressor
Inducible: something inactivates repressor which then induces transcription
201
Negative Repressible Operon
- Transcription normally takes place and must be turned off
- inactive in the absence of a co-factor
- co-repressor binds to the repressor which allows it to bind to the operator and inhibit transcription
Negative: regulator protein is an inhibitor or repressor
Repressible: Something activates the repressor which. then represses transcription
202
Positive Control - Gene expression
- regulatory protein is an activator
- Activator binds operator and induces txn
- Binds just upstream of the promoter and enhances the binding of RNA polymerase to the promoter
203
Describe the lac operon
Negative inducible:
- regulator is an inhibitor
- allolactose inactivates inhibitor
Positive Control:
- CAP+cAMP enhances transcription
204
what is the role of permase (lacY)
To transport lactose across the bacterial cell membrane
205
Role of b-galactosidase (lacZ)
catalyzes the breakdown of lactose to glucose and galactose
also converts lactose into allolactose
206
Cis regulation of expression
control of gene expression of genes only on the same piece of DNA
207
Trans regulation of expression
Control gene expression on other DNA molecules
208
non functional mustations are designted by
-
209
Catabolite Repression
Bacteria will actively turn off the other metobolic pathways when glucose is present
209
Positive Control
Positive glucose metabolism results in repression of other pathways
209
levels of cAMP are ___ proportional to glucose
inversely
glucose low = cAMP high
