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BB 492 > Exam 1 > Flashcards

Exam 1 Flashcards

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1
Q

SOS Response

A
  • Prokaryotes
  • Last resort induction of genes involved in DNA repair
  • LexA, RecA
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2
Q

LexA

A

Transcriptional repressor that keeps SOS genes turned off under normal conditions.

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3
Q

RecA

A

Protein that binds to ssDNA and stimulates cleavage of LexA.

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4
Q

NHEJR

A
  • Fixes double-stranded breaks
  • High risk of Indels
  • Ku70/80 heterodimer, TdT
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5
Q

Ku70/80

A

Protein that associates with the severed ends and acts as a docking site during NHEJR.

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6
Q

Terminal Deoxynucleotidyl Transferase (TdT)

A

A DNA polymerase that adds non-templated nucleotides during NHEJR.

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7
Q

Nucleotide Excision Repair

A
  • Recognizes DNA helix distortions (adducts, thymine dimers, etc.)
  • Exinuclease
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8
Q

Exinuclease

A

Excision endonuclease that hydrolyzes phosphodiester bonds on either side of the lesion during NER. Segment is removed (helicase), then gap repaired.

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9
Q

Photoreactivation

A
  • Form of direct reversal
  • Visible light energy breaks damaged DNA structure, restoring original pyrimidines.
  • Assc. with UV damage
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10
Q

AGT

A
  • Form of direct reversal
  • Enzyme that removes an alkyl group on an alkylated base
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11
Q

AlkB-related Dioxygenases

A
  • Form of direct reversal
  • Enzyme that modifies an alkyl group on an alkylated base.
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12
Q

Direct Reversal

A
  • Damage is directly fixed, without cutting the backbone or excising nucleotides.
  • Rare
  • Photoreactivation, AGT, AlkB-related dioxygenases
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13
Q

Base Excision Repair

A
  • Fixes damage to specific nucleobases
  • Single base lesions, single strand breaks
  • DNA glycosylases, abasic site, APE
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14
Q

DNA Glycosylase

A
  • Scan DNA for lesion they repair (recognize conformational changes)
  • Binds to base, flips base out of the backbone and into active site
  • Hydrolyzes the N-glycosidic bond, leaving abasic site
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15
Q

AP Endonuclease (APE)

A
  • Performs lesion removal during BER
  • Pinches strand so that lesion enters exonuclease site
  • Cleaves backbone (endonuclease), leaving 3’ -OH and 5’ dRP
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16
Q

Pol Beta

A

Trims out dRP in short-patch BER, adds correct dNTP

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17
Q

DNA Ligase III

A

Ligates gap left from short-patch BER

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18
Q

Which form of BER is preferred when the cell isn’t proliferating?

A

Short-patch (replication machinery is not available for long-patch)

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19
Q

Long-Patch BER

A
  • Pol delta or Pol epsilon/PCNA recognizes and drives through lesion
  • FEN1 + PCNA excises flap containing AP site
  • DNA ligase I recognizes and repairs
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20
Q

DNA Ligase I

A

Ligates gap left from long-patch BER

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21
Q

Mismatch Repair

A
  • Corrects mistakenly incorporated bases and indels
  • Methylation distinguishes in prokaryotes
  • Different mechanisms between the domains
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22
Q

MutS

A

Protein that recognizes and forms a dimer at the mismatch site during MMR

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23
Q

MutL

A

Protein that binds to MutL (when it dimerizes), forming an active complex during MMR

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24
Q

MutH

A

Cleaves unmethylated strand on the 5’ site of the GATC (has to be paired to hemimethylated GATC) during MMR

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25
DAM Methylase
E. coli protein that methylates all adenines in GATC sequences
26
Eukaryotic MutS homologs
MSH2, MSH3, MSH6
27
Eukaryotic MutL homologs
4 heterodimeric homologs
28
Eukaryotic MutH homolog
None known
29
What is involved in identification of strands during eukaryotic MMR?
- 5' end of Okazaki fragments - Single strand breaks
30
3 Types of mutation
- Point Mutation - Frameshift Mutations (Indels) - Single-strand or double-strand breaks
31
Example of Transition
Methylating agents (alkylating agents, cigarette smoke, S-AdoMet)
32
Example of Transversion
Cytosine Deamination (C deaminates to U, U pairs with A, DNA repairs U by swapping it with T) resulting in change from CG to AT
33
Mutagens associated with base modifications and single-strand breaks
- ROS - Alkylating Agents - X-rays - Spontaneous changes (RAXS)
34
Mutagens associated with replication errors
- Mismatched base insertion - Indels - Topo II adducts
35
Mutagens associated with bulky stable adducts
- UV damage - Mutagens from food - Intercalating agents
36
Mutagens associated with inter-strand adducts and double-strand breaks
- Chemo - Xrays - Ionizing radiation (CXI, all medical)
37
What phenomenon is associated with 1/3 of all single site mutations?
Base deamination
38
What causes indels?
- Polymerase "slippage" off template - Whether it is insertion or deletion depends on which strand loops out
39
Topoisomerase "Trapping"
- Stabilizes Topo II cleavage complex (still covalently bound to 5' phosphotyrosyl linkage) - Misalignment of 5' -OH DNA end - Topo II cuts DNA, but fails to reseal it
40
Camptothecin
- Anticancer - Inhibits topo I by trapping (topo-DNA complex stabilized)
41
Ciprofloxicin
- Broad-spectrum antibiotic - Impacts Tpo II
42
Doxorubicin
- Chemo - Antibiotic - Traps topo II (stabilizes topo-DNA complex)
43
Etoposide
- Chemo - Traps topo II
44
Novobiocin
- Antibiotic (gram pos) - Inhibits GyrB via competitive inhibition against ATP binding
45
3 causes of replication errors
- Altered dNTP pools - "Low fidelity" polymerases - "Failure to remove an incorrect base
46
What are four factors that help maintain genomic sequences?
- Watson-Crick base pairing - DNA polymerase proofreading - Packaging - Telomeres
47
What supercoiling does Topo II relieve in the context of the replication fork?
Both positive and negative
48
What supercoiling does Topo I relieve in the context of the replication fork?
Positive only
49
Telomeres
Tandem repeats of a short, G-rich sequence on the 3' ends of eukaryotes.
50
Tert
The reverse transcriptase component of telomerase
51
TERC
The ribonucleotide segment of telomerase. Is complementary to the telomeric sequence and acts as a template for extending the 3' end of the strand.
52
What are the two components of telomerase?
Tert, TERC
53
G-Quartets
Aggregations of guanine-rich sequences, caused by telomeres
54
Eukaryotic SSBP homolog
RPA
55
Pol Delta
- Polymerase used for most of the lagging strand synthesis in eukaryotes - Associates with PCNA (which then associates with FEN1) to get rid of the primer
56
Pol Epsilon
- Polymerase used for most of the leading strand synthesis in eukaryotes
57
Eukaryotic Beta Clamp Homolog
PCNA
58
Pol Alpha
Associates with primase to lay down RNA primer in eukaryotes
59
Pre-Licensing
Involves pre-replication complex, ORC
60
Eukaryotic DnaA homolog
ORC (Origin recognition complex)
61
cdc6
Protein that binds to ORC following conformatoin change
62
Eukaryotic DnaB homolog
MCM ("licensing proteins")
63
MCM
Protein hexamers that use ATP hydrolysis to move and unwind DNA in eukaryotes
64
cdc45
Protein that is required for activation of the pre-recognition complex in eukaryotes
65
GINS
- Protein that is required to activate the pre-recognition complex in eukaryotes - Binds ssDNA and recruits primase/pol alpha
66
Degradation of what protein prevents continuous licensing?
cdc6 (removed/degraded when MCM hexamers begin separating dsDNA)
67
Primase/Pol Alpha Complex
- Lays down primer in eukaryotes - Primase binds to GINS - Pol alpha binds to primase
68
What protein controls the accumulation and ability of DNA replication proteins to perform DNA synthesis?
Cyclins/CDKs
69
What molecules control cell cycle checkpoints?
Cyclins and CDKs
70
During what phase of the cell cycle is DNA synthesis initiated?
Late G1
71
3 main ways that eukaryotic replication differs from prokaryotic
- Requires multiple DNA polymerases - Preprelication complexes assemble (during cell cycle, multiple origins) - Telomeres
72
Topo IV
Topoisomerase that catalyzes the separation of catenanes during termination of prokaryotic replication
73
Tus
Protein that binds to the Ter site in a specific direction (terminus utilization sequence)
74
Ter
Sequence opposite the OriC that serves as a binding site for the Tus protein.
75
Tus-Ter Complex
Acts as a roadblock that arrests the replication fork in one direction.
76
DNA Pol I
Removes RNA primers via "nick translation" during prokaryotic replication - RNs to dRNs, moves nick to 3' end of new DNA
77
2 distinguishing features of DNA ligase
- Hydrophobic pocket for NAD or ATP - Cleft for DNA binding
78
DNA Ligase Mechanism
- Critical lysine adenylated - Nicked DNA moved closer to adenylated lysine. - Adenyl group transferred to 5' phosphate of nick - 3' -OH attacks 5' phosphate to close the gap
79
Pol III Holoenzyme
Performs primary DNA synthesis during replication in prokaryotes
80
Pol III holoenzyme components
- 2 catalytic cores, each with template strand - Linked gamma complex ("clamp loader") - Beta clamp (helps cores stay attached to DNA)
81
Clamp loading process
- Beta clamp and 3 ATPs bind to clamp loader - ATP binding forces the beta clamp to open, DNA enters - ATP hydrolysis causes the clamp to close around the DNA
82
Replisome
The machinery that carries out DNA replication in prokaryotes (includes pol III holoenzyme)
83
Proteins that are not part of the replisome, but still needed for elongation in eukaryotes
- DNA gyrase - SSBPs
84
How is eukaryotic synthesis coordinated?
Pol III directly coordinates with DnaB and DnaG
85
Primosome
The complex that initiates synthesis of the RNA primer in prokaryotes.
86
DnaG (Primase)
DNA-dependent RNA polymerase that associates with DnaB on the separated strands during prokaryotic synthesis.
87
Where does DNA pol III add dNTPs?
To the free 3' -OH of the primer
88
Primer formation on prokaryotic lagging strands
- Primosome has to initiate each Okazaki fragment - Primosome moves in 5' to 3' direction on template, so has to reverse to lay down primer
89
How is helicase moved into the replication fork?
Conformational changes induced by ATP hydrolysis
90
DNA Unwinding Element (DUE)
An AT-rich sequence near the OriC in prokaryotes that is prone to unwinding. Creates binding sites for replication machinery.
91
What helps with coordination in prokaryotic polymerases?
Mg2+ coordinates Asp residues and phosphate groups on dNTP
92
How are nucleotides added during replication?
Nucleophilic attack of growing chain's 3' -OH on alpha-phosphate of incoming dNTP
93
Class II Topoisomerases
- Multimeric - ATP, Mg2+ Dependent - Changes Lk by 2 - Can relax negative and positive supercoils - Strand Passage
94
Class I Topoisomerases
- Monomeric - Change Lk by 1 - Mechanism depends on type
95
Topo IA
- Strand-Passage Mechanism - Phosphotyrosine bond on 5' terminal phosphoryl - Eukaryotes and prokaryotes - Relaxes negative SCs
96
Topo IB
- Controlled Rotation Mechanism - Phosphotyrosine bond on 3' terminal phosphoryl - Eukaryotes only - Relaxes positive and negative SCs
97
DNA Gyrase
A form of Topo II found in E. coli that can introduce negative supercoils
98
Heterochromatin
Densely packed, less active
99
Euchromatin
Less densely packed, active
100
Nucleosome
DNA + Histones (fundamental unit of chromatin)
101
Chromatin
Duplex DNA + All Proteins
102
5 histone classes
H1 H2A H2B H3 H4
103
What protein facilitates coiling of DNA around histones?
Topoisomerase (relieves strain from the negative supercoiling)
104
Surface residues associated with histones
Lys & Arg
105
Relaxed DNA maintains the standard rise of ___ bp per turn.
10.5
106
Negative supercoiling
Twisting against the helical conformation (left-handed direction)
107
Positive supercoiling
Normal conformation of DNA is twisted even tighter
108
Linking Number
The number of times that one strand wraps around another (topological).
109
Linking number equation
Lk = Tw + Wr
110
Tw = ?
bp/10.5
111
Wr = ?
0 in relaxed DNA
112
Delta Lk equation
Delta Lk = Lk - Lk(0)
113
2 criteria needed to supercoil DNA
- No breaks in either strand - Strands have Lk of 1
114
Superhelical density equation
Omega = Wr/Tw (compares supercoiling)
115
Base conformation in A-DNA and B-DNA
Anti
116
Conditions favoring DNA denaturation
- High temperature - Low salt concentrations - High pH
117
Conditions favoring DNA reannealing
- Low temperature - High salt concentrations - Neutral pH
118
B-DNA
- "Classic" form - Right-handed winding - Most thermodynamically stable
119
A-DNA
- Rare, only seen in dehydrated state - 20* rotation of helix relative to B-DNA - Deep major groove, flat minor groove
120
Z-DNA
- Transient - Left-handed helix - Greater distance between base pairs
121
What base component is the hydrogen donor?
Amino group ("Amino adds")
122
What base component is the hydrogen acceptor?
O and ring N ("Take ON")
123
How does ribose differ from deoxyribose?
Ribose has 2' -OH
124
5 categories of nucleotides
- Coenzymes - Regulatory molecules - Information - Macromolecule metabolism - Energy currency (CRIME)
125
5 routes of information flow described by the central dogma
- Replication - RNA Replication - Transcription - Reverse Transcription - Translation

BB 492 (4 decks)

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