M2M Unit 1 Flashcards
(309 cards)
1
Q
function of aminoacyl tRNA synthetase
A
put the correct AA on the correct tRNA
2
Q
during translation, the mRNA is decoded by what process
A
the anticodon loop of a cognate tRNA pairs with the 3 nucleotide in mRNA
3
Q
basic elongation cycle: 3 truths and 1 false
A
tRNAs enter into the A site and leave from the E site
peptide bond formation results in the transfer of the growing peptide chain from the P site tRNA to the A site tRNA
translocation occurs after peptide bond is formed
FALSE TRANSLOCATION REQUIRES ATP HYDROLYSIS
4
Q
homologous recombination depends on:
A
the availability of the sister chromatid
5
Q
during normal ds break repair:
A
the repaired sequence is indistinguishable from the sequence prior to the break if homologous recombination is used
6
Q
the decision to use NHEJ instead of HR depends on:
A
blocking resection of the DNA ends
the activity of ligase 4
time in the cell cycle
NOT BRCA2
7
Q
microRNAs play a role in:
A
translational repression
8
Q
the mRNA cap serves important roles in splicing, 3’ processing, nuclear export, and translation. the 1st enzymatic step in the addition of the cap to the newly synthesized mRNA is:
A
triphosphatase
9
Q
the proliferating cell nuclear antigen (PCNA) is a key component of the eukaryotic replisome. What function does it perform?
A
acts as a clamp which encircles the DNA and binds the replisome to affect processive synthesis of DNA
10
Q
nuclear receptors such as estrogen receptor are transcription factors that mediate expression of hormone-regulated genes. these nuclear receptors are proteins which contain 3 identifiable domains:
A
transactivation domain
DNA binding domain
hormone binding domain
11
Q
what nuclear receptor domain contains a Zn finger structure critical for steroid receptor function?
A
DNA binding domain
12
Q
the fidelity of DNA replication occurring at the DNA replication fork comes from the: 3 things
A
inherent accuracy of the active site of the replicative DNA polymerase to select the correct incoming dNTP
the 3’-5’ proofreading exonuclease of the DNA polymerase to excise an incorrectly inserted nucleotide added to the 3’ OH terminus
a DNA repair activity that follows the replication fork
13
Q
what type of DNA repair activity follows the replication fork?
A
mismatch DNA repair
14
Q
Cockayne syndrome, Xeroderma Pigmentosum, and Trichothiodystrophy are all deficient in which DNA repair pathway?
A
nucleotide excision repair NER
15
Q
during the initial phase of translation in bac, what guides binding of the mRNA to the 30S subunit of the ribosome?
A
pairing of the shine-delgarno sequence with the 16S rRNA
16
Q
during splicing of the pre-mRNA, how is the 5’ slice site recognized?
A
base pairing of the U1 snRNA
17
Q
what type of atomic interaction plays the central role in formation and stability of the alpha helix and beta sheet structures?
A
hydrogen bonding
18
Q
which AAs are used to create turns in secondary structures?
A
proline and glycine
19
Q
beta-amyloid protein production occurs as a result of which enzymatic cleavage sequence?
A
beta-secretase followed by gamma-secretase action on beta-amyloid precursor protein
20
Q
how does Swi/Snf complex remodel chromatin?
A
uses HTP hydrolysis to break histone-DNA contacts and move histone octamer down DNA
21
Q
what is the major role of binding of the TATA box binding protein to the TATA box?
A
helps direct assembly of the preinitiation complex
22
Q
in the Michaelis-Menton description of kinetics, Kcat describes:
A
the number of substrate molecs converted to product at saturating substrate conc’s
23
Q
which bond is broken during ATP hydrolysis?
A
beta-gamma phosphoanhydride bond
24
Q
what intracellular conc of ADP will provide the most negative delta G for hydrolysis of the ATP?
A
lowest ADP conc (LaChatlier’s principle)
25
the RNA pol2 transciption factor IIH includes:
XPB protein
BPD protein
Cdk7, a cyclin-dependent protein kinase
NOT TATA BINDING BOX PROTEIN
26
what happens during splicing of euk mRNA
an intron is removed and 2 exons are joined
27
what does a spliceosome recognize
GU at the beginning of an intron
AG at the end of an intron
A branch point residue
NOT 5' UTR
28
normal peptide with 100 AAs, has a mutation, now it only has 20. 1-10 are normal, but 11-20 are different. what happened?
nucleotide deletion in the codon for AA 11
29
you prepare a solution of glucose in water at standard conditions. why doesn't any of it break down?
steps in metabolism of glucose require Ea
30
high Energy bonds (4)
```
ATP (phosphoanhydride)
phosphocreatinine P-N
acetyl CoA
phosphophenolpyruvate C-O-P
NOT PHOSPHOSERINE
```
31
what reagent is used in Western blot to help detect a specific macromolec that has been transferred from the electrophoretic gel to the blotting membrane?
antibody
32
exome sequencing experiments are designed to interrogate which parts of the genome?
protein-coding regions
33
patients with Lynch syndrome (hereditary nonpolyposis colon cancer syndrome) have defects in which DNA repair pathway?
mismatch DNA repair
34
3 true statements about chromatin remodeling:
the ability of transcription factors to bind DNA can be affected by chromatin structure
- the SWI/SNF remodeling activity is an ATP-dependent ATPase that helps disrupt histone octomers
- HATs are co-activators
- NOT N-TERMINI OF HISTONES ARE RICH IN ARGININES (should be lysine)
35
what's different between bac and euk RNA polymerases?
RNA polymerase is modified by phosphorylation of the CTD when it clears the promoter
36
3 similarities between bac and euk transcription polymerases
5' end of primary transcript is a triphosphate
- isomerization results in formation of an open complex at the promoter
- when RNA polymerase encounters a block, an RNA endonuclease activity is stimulated
- RNA synthesis is completely processive
37
what role does eIF4E play?
functions as a cap binding protein
38
what can accurately describe G1 phase
cell division and growth are coordinated in G1
39
during prokaryotic translation, recognition of the shine delgarno sequence in mRNA allows:
placement of the correct AUG at the P site
40
what does Avastin do to treat tumors?
inhibits formation of new blood vessels induced by the vascular endothelial growth factor (VEGF)
41
what kind of molec is Avastin?
humon monoclonal antibody
42
what enzyme is used for next generation DNA sequencing?
DNA polymerase
43
what is most easily analyzed by short read DNA sequencing
genotype
44
energy used to catalyze DNA joining by ligase comes from
ATP, to promote high-E phosphodiester formation
45
4 common types of post-translational protein modification
phosphorylation of threonine
methylation of Lysine
addition of sugars to asparagines
acetylation of lysine
46
alzheimer's disease is thought to be caused by
misfolded proteins
47
control of transciption is combinatorial, meaning:
few transcription factors come together in different ways to control transcription of many genes
48
normal gene expression can be controlled by:
mRNA export from nucleus
| efficiency in translation
49
sensor proteins in the DNA checkpoint pathway:
bind DNA with blocked replication forks
50
how do human somatic cells maintain their size
cells arrest in G1 phase until they are big enough
51
where does the phosphate attach for DNA and RNA building blocks in the ribose?
5' carbon
52
short read and long read reading lengths:
100bp and 10,000 bp
53
Laws of Thermodynamics
1st- E is conserved
| 2nd- entropy is always increasing
54
7 forms of E
```
Kinetic
Potential
radiant
thermal
mechanical
electric
chem
```
55
thermodynamic equations
```
deltaG0= -RTlnKeq
deltaG= deltaH-TdeltaS
deltaG= -nFE (redox)
```
56
Keq related to deltaG
deltaG is negative when Keq is >1
| deltaG is 0 when Keq is 1
57
how can you make a nonspontaneous rxn favorable?
couple it with a spontaneous rxn
| standard free E changes are additive
58
2 High E compounds:
lipids
carbs
both have excess electrons
59
Purine vs Pyrimidine
Pure As Gold AG
| CUT the Py
60
deoxyribose vs ribose
2' -H or 2' -OH
61
nucleoside vs nucleotide
```
nucleoside= sugar + base
nucleotide= sugar, base, + P
```
62
relative solubility rule
phosphate > nucleotide > nucleoside > pyrimidine > purine
63
Gout and Lesch-Nyhan Disease are related to:
excess insoluble purine build up in tissues and kidneys
64
chemistry of phosphodiester bonds:
3' -OH on ribose bonds with 5' phosphate
| ends of DNA/RNA chains will either have an exposed 3' -OH or an exposed 5' Phosphate
65
Avery, McCloud, and McCarty's exp:
est DNA as genetic material through pneumococcus exp:
| S strain killed, R strain didn't; DNA from heat-killed S cultured with R killed
66
Franklin and Wilkins exp:
x-ray diffraction suggesting helical structure
67
Watson and Crick:
discover definitive double helix structure
68
Chargaff's Rule:
suggested base pairing
| ratios of G:C and A:T were equal
69
3D model for DNA:
right-handed, antiparallel, double stranded helix
hydrophilic sugar phosphate backbone on outside
hydrophobic base pairing on inside
major and minor grooves
about 10 bp per helical turn
70
how are electrostatic repulsions neutralized in DNA backbone?
- positively charged species in cell (Mg)
- base pair linkages offer stability
- adj base pair stacking, offering delocalization
71
what does salt, pH, length, and GC content do to DNA stability
salt inc stability and Tm
pH extremes dec stability (alter base ionization and H bonding)
length inc stability
high GC content inc stability (3 H bonds- more delocalization)
72
linear vs circular DNA
linear: large, segments can become supercoiled
Circular: shorter; bound to itself, so no supercoiling problem
73
5 chemical modifications of DNA bases
```
methylation
deamination
depurination
UV light
alkylating agents
```
74
DNA methylation
typically CpG--> Met-CpG
catalyzed by DNA methyltransferase
80-90% human CpG sites are methylated
75
CpG island
GC rich area without methylation
| less methylation = more transcriptional activity
76
DNA deamination
C loses an amine --> U
| potentially changes CG pair to a TA pair
77
DNA depurination
hydrolysis of N-glycosidic bond to lose purine base
-OH replaces purine loss
weakens backbone
78
DNA UV light interaction
UV light can dimerize adjacent Thymines
| distorts DNA helix and blocks replication enzymes
79
UV damage repaired by:
nucleotide exicision repair NER
80
DNA alkylating agents
nucleophilic attack of bases on nucleotides
| mustard gas, cisplatin
81
DNA polymerization and nuceloside analogs used for drug therapy
nucleoside analogs block replication of virally infected cells by being incorporated into the replicating chain
analogs make DNA chains nonfunctional
82
when are more and less specific nucleoside analogs used, respectively
more specific- used against retroviruses
| less specific- used against cancer
83
4 methods to attack DNA metabolism:
block synthesis of precursors
intercalation
covalently bind bp's
attack topoisomerases
84
using DNA probe and Tm for diagnostic techniques:
complementary DNA strands have high Tm
| use a probe of known sequence against unknown sequence to see if Tm is same or lower (mutation)
85
3 classes of RNA in human cell
Structural
regulatory
information-containing
86
4- structural RNA's and func's
rRNA- make up ribosomes
tRNA- move RNA
snRNA and snoRNA- splicing and other in-cell modifications
87
2 regulator RNA's and func
miRNA and siRNA- downregulate gene expression
88
1 info-containing RNA and func
mRNA- to be translated into proteins
89
how does puromycin mimic amino-acyl tRNA to terminate translation
puromycin is an antibiotic; mimics the acceptor 3' end of tRNA
it covalently attaches to polypep chain and prevents completion of translation
90
origin of replication
specific sequence recognized by binding proteins
usually multiple short repeats w/ AT rich streak
1 in prokaryotes, 100s per chromosome
91
origin binding proteins
bind to the origin and become part of the complex
| recruit Pol3
92
helicases
catalyze breakage of H bonds to unwind helix
93
SSB protein
bind to melted strands of DNA to prevent re-annealing
| esp important for okazaki fragments
94
primase
enzyme that catalyze the the addition of RNA primer to being replication
95
DNA Pol1
DISTRIBUTIVE- replaces RNA primers using:
DNA polymerase, 3-5' exonuclease activity, and 5-3' exonuclease activity
NO binding clamp- so it's slow and distributive
96
DNA Pol3
PROCESSIVE- synthesizes DNA strand from its compliment
| bound tightly to DNA via sliding clamp- works fast and is processive
97
common to Pol1 and Pol3
proofreading activity AKA 3-5' exonuclease activity
98
DNA ligase
enzyme responsible for sealing okazaki fragments once RNA primers ahve been replaced by Pol1
99
Telomere
sequence at end of chromosomes; large repeated segments
progressively shorter w/ each replication
cell becomes destroyed when telomeres become too short
100
telomerase-
enzyme ensures telomeres never shorten in immortal cells (germ cells)
act as reverse transcriptases on DNA ends
REPRESSED in somatic cells
DE-REPRESSED in cancer cells
101
topoisomerase/gyrase
enzyme responsible for relieving torsional strain in DNA helix AHEAD of replication fork
(gyrase is specific to prokaryotes)
102
reverse transcriptase-
enzyme responsible for copying INTO DNA usually from RNA
| can be endogenous or exogenous
103
how DNA polymerase creates phosphodiester bond
breaks off phosphodiester bond from dNTP and uses liberated E to bind remaining P group to OH group on previous nucleotide in the chain
104
Does DNA polymerase require an RNA primer?
YES
105
DNA synthesis leading vs lagging strand
leading: orign binding proteins bind; DNA melted; topoisomerases relieve tension; Pol3 elongates DNA; 2 semiconserved strands are annealed
lagging: fragments are sealed together with DNA ligase
106
"End Replication Problem"
lagging strand can't be synthesized all the way because RNA primer can't attach past the end of the DNA
leads to shortened telomeres
telomerase maintains chromosomal ends in germ and cancerous cells
107
3 criteria for cell to become cancerous:
mutation/mismatch in a gene with proliferation;
cell can't fix the problem
self-destruction is repressed/inhibited
108
xeroderma pigmentosum and cockayne syndrome are 2:
inheritable diseases from defective DNA repair
109
4 sources of DNA damage:
thymine dimers
uracils in DNA
bulky chemical adducts
double stranded breaks
110
thymine dimers
NER
UV causes adj thymines to bond, causing kinks
can cause Pol3 to fall of and Pol2 to take over (lots of errors)
111
uracils in DNA
BER
cytosine deamination --> uracil
can cause replication, transcription, and recognition problems
112
bulky chemical adducts in DNA:
NER
| similar to thymine dimers, but caused by toxic large molecs binding to DNA bases
113
double-stranded breaks in DNA:
HR or NHEJ
| can potentially lose half a chromosome
114
process of mismatch repair
mismatch recognized shortly after synthesis
endonucleases clip on either side
exonuclease and helicase excise problem
DNA Pol3 replaces w/ correct sequence (sealed by ligase)
115
mismatch reconition in E coli vs humans
E coli- old strand is methylated, so endonucleases clip the un-methylated strand
humans- unknown mech
116
nucleotide excision repair NER
repairs more overt problems that alter helix
| recognize, clip by endonucleases, excise affected part, replace by Pol1, ligate
117
NER recognition pathway needs:
transcription factor TF2H
118
2 kinds of NER
transcription-coupled NER: w/in gene being actively transcribed
global genome NER: not w/in gene actively being transcribed
119
how does DNA replication continue with lesions? 2 ways
lesion bypass polymerization: usually occurs when cell can't fix all the thymine dimers
bypass polymerases: attach when DNA Pol3 is stopped and NER can't occur; they add nucleotides without proofreading (high error rate)
120
DNA damage checkpoint maintaining genome stability
cell cycle is paused if damage is sensed; repair machinery is up-regulated; 2 kinds of protein kinases (ATR and ATM) amplify signal to phosphorylate Chk1 and Chk2 for repair, arrest, or death. Chk1 and 2 phosphorylate p53, which causes cell cycle arrest or death. Chk1 and 2 kinases ensure problems have been fixed;
121
mutations in DNA damage checkpoints
lead to genomic instability- cell can't regulate itself and may proliferate into cancer
this checkpoint is first step in guarding against cancer
122
functions of ATR and ATM
both are protein kinases in DNA damage checkpoint
ATR is for stalled forks
ATM is for double stranded breaks
123
RNA polymerase reaction
unidirectional
catalyzes phosphodiester formation in 5' to 3' direction based on 3' to 5' template
need tripphosphate nucleotide for spontaneous rxn
124
5 steps in transcription cycle common to bac and euk RNA polymerases
INITIATION
1- RNA polymerase binds to promotor sequence on DNA in closed complex
2- polymerase melts DNA near transcription start site- form transcription bubble
3- polymerase catalyzes phosphodiester bond of two initial rNTPs
ELONGATION
4- polymerase adv 3' to 5' on template strand, melting DNA and linking rNTPs
TERMINATION
5-polymerase releases completed RNA and dissociates from DNA at transcription stop site
125
4 cellular RNA polymerases and func's
RNA pol1- makes rRNA
RNA pol2- makes mRNA, snRNA, miRNA
RNA pol3- makes tRNA, lncRNA
mitochondrial RNApol- makes mito RNA
126
promoter:
where RNA polymerase binds
| sequence of DNA upstream of the transcription start site that positively affects expression of gene
127
4 consensus elements of promoter and func
TATA box- 30bp upstream TATA sequence
initiator- +1 in some but not all euk genes
promoter proximal elets- promoter DNA sequences 30-1000 bp upstream
enhancer elets- even farther upstream; acts through DNA looping
128
TATA mutation consequences
TATA box binding protein helps assemble pre-initiation complex at the promoter, so a mutation would cause reduced expression of gene
ie beta-thalassemia with B-hemoglobin
129
how does alpha-amanitin block transcription
toxic sub. found in death cap mushrooms; inhibits RNA Pol2; BINDING ITS BRIDGE SUBSTRUCTURE translocation down the DNA chain can't happen
130
how does rifampicin block transcription
broad-spectrum antibiotic; acts by binding the beta subunit of bac RNA polymerase, PLUGS UP THE EXIT CHAMBER so elongation can't occur
131
4 components in RNA polymerase 2 pre-initiation complex
RNA pol2
general transcription factors (TF2_)
promoter DNA
mediator
132
mutations in TF2H subunit syndromes:
TF2H is a transcription factor that also repairs DNA damge
cockayne's syndrome and trichothiodystrophy= problems w/ NER
xeroderma pigmentosum- light sensitivity; cancer susceptibility; abnormal neuro; unscheduled DNA synthesis
133
3 major ways most pre-mRNA's are processed:
capping- replace 5'triPhosphate w/ 7-methylguanosine
splicing- excision of introns and desgementation of exons
cleavage/polyA- cleavage at 3' end past consensus sequence and polyA of cleaved site
NOTE these take place while RNA is still being made
134
pre-mRNA vs mature mRNA
pre: 3' phosphate intact; has introns; 3' end is unmodified
mature: 7-methylguanosine cap; spliced out introns; polyA tail added to 3' end
135
4 funcs of 5' mRNA cap
5' end is resistant to exonucleases
helps with splicing and processing through cap-binding complex
translation factor eIF4E recongizes cap for ribosome transport
signals for mRNA to degrade when cap is removed
136
3 rxns required to add a 5' mRNA cap to pre-mRNA
1- cut off last Phosphate at the 5' end
2- add GTP backwards via guanylyl transferase (losing 2 phosphate groups)
3- methylate 7-position of guanosine cap via SAM (s-adenosyl methionine)
137
conserved intron and consensus sequence in polyA tail
5' intron: GU
3' intron: AG
consensus sequence: AAUAAA
138
how does alternative splicing permit multiple proteins to be produced by splicing defects
5' splice site (Identified by snRNA) is a part of the intron, so if it's accidentally made unrecognizable then it will be translated and the mRNA will continue on, creating a significantly different protein
139
disorders caused by splicing defects:
Marfan's syndrome- disruption of the fibrillin gene; pts are tall and prone to aneurysms
abnormal splicing of CD44- predictor of tumor metastasis
140
func of U1snRNA, U2snRNA, and U2AFsnRNA in splicing
U1- binds to the GU 5' splice site of introns
U2- binds to A branch point
U2AF- binds to AG 3' splice site in introns
141
what does Lariat splicing mech do
uses the U1, U2, and U2AF splicing mech's to link 2 exons and excise the intron complex
142
what is 5' UTR and 3' UTR
5'- region between +1 and start codon on processed mRNA
| 3'- between stop codon until end of transcript, incl consensus seq and polyA tail
143
2 rxns to make mature 3' mRNA
1-recognize consensus seq at pre-mRNA's 3' end and cleave soon after sequence
2- polyA of free -OH at 3' end
144
how is 3' pre-mRNA end processing related to transcription termination
cleavage and polyA continue during RNA polymerase, it could soon fall off or could continue making mRNA
polyA tail seems necessary for RNA polymerase complex to detach itself and terminate
145
2 func's of mRNA polyA tail
protection from degradation
| export mRNA from nucleus
146
alternative polyA sites used to make more than 1 protein in a single gene
2 forms of immunoglobulin M exist because transcription can continue past 1st polyA site and end on 2nd polyA site (heavy and light chains)
147
what are DNA control elets
```
DNA elets that act locally
transcription factors bind to regulated gene expression
TATA box/initiator sequence
promotor proximal elet
enhancers
```
148
what are DNA transcription factors
proteins encoded by 1 gene that act on other genes to regulate their transcription
can activate/repress many genes
149
promotor proximal elet vs enhancer
PPE- usually w/in 200 bps upstream of start site; about 20 bp long; bound by transcription factors that regulate transcription
enhancer- much farther upstream/downstream than promoters; similar size and func to promoters
150
beta-thalassemia:
mild inherited anemia due to promoter mutation of b-globin gene, so less b-globin is produced
151
gamma-delta-beta thalassemia
more serious anemia caused by a deletion in the control region for transcription of all globin genes, so loss of globin translation
152
hemophilia b leydon:
x-linked disease affecting clotting
problem with promoter region of clotting protein gene
tends to improve partially at puberty
153
fragile x syndrome
x-linked
mental retardatyion, atypical face development w/ enlarged testicles
caused by expansion of CGG count upstream of a gene that causes high rate of methylation and transcriptional silencing of that gene
154
role of transcriptional activators and repressors
bind to DNA control elements or to other factors bound to control elements
increase/decrease rate of transcription of that gene
155
2 classes of transcriptional activators/repressors
sequence specific binding proteins SSB- bind to control elets in DNA via alpha-helix insertion in major groove
co-factors: bind to SSBs- can increase or decrease transcription efficacy
156
2 domains of SSBs
1st- DNA binding domain; binds to DNA target; highly structured and conserved
2nd- activation of domain; recruit other proteins (cofactors or GTFs) to bind and affect transcription
157
4 families of SSBs, based on tertiary structure differences
homeodomain
Zn finger
basic leucine zipper
basic helix-loop-helix
158
homeodomain proteins
SSB
| helix-turn-helix structure; affect many genes at once
159
zinc finger protein
largest family of SSB
includes estrogen and androgen receptors
finger made of 2 antiparallel beta sheets and alpha helix, held together by Zn ion
finger binds w/ DNA
160
basic leucine zipper protein
SSB- chop sticks
hydrophobic residues
dimerizes to bind DNA
161
basic helix loop helix protein
SSB
| muscle group
162
craniosynostosis
mutation in SSB- homeodomain that upregulates gene
| premature skull closure
163
androgen sensitivity syndrome
mutation in SSB- Zn finger androgen receptor
downregulates transcription of genes controlled by male androgens
feminization
164
Waardenburg syndrome
mutation in SSB- basic helix loop helix protein
| deafness, pigmentation defects
165
how is cominatorial control used as a mech for controlling gene expression
many SSB proteins can dimerize, to make many DNA binding sequences within the same family of SSB's (Zn finger w/ Zn finger)
166
how does chromatin structure affect transcriptional control
DNA-dependent ATPases: disrupt histone octamers, opening chromatin and exposing it for binding via hydrolysis (Swi/Snf complex)
167
2 factors that irreversibly modify histones through acetylation of N-terminus
HATs- acetylators, co-activators
| HDACs- deacetylators, co-repressors
168
current theory on HATs
pattern of histone acetylation recruits co-factors to affect increased transcription rather than directly affecting it themselves by charge
169
leukemia and histone activity
haematopoietic (blood)
| chromosomal translocations over- activating fusion proteins that alter HDAC/HAT activity
170
rubinstein-taybi syndrome and Histone activity
growth and mental retardation; broad thumbs and toes; craniofacial dysmorphism
mutations in one copy of CREB binding protein gene, a HAT important in development
171
activators/repressors interaction with general transcription machinery vs chromatin
bind to GTFs or RNA Pol2 complex to influence initiation or elongation of primary transcript
regulate accessibility of DNA on chromatin to Pol2 transcription apparatus (acetylation, phosphorylation, methylation, ubiquination)
172
how is specificity achieved with transcriptional regulators
specificity depends on binding with specific DNA control elets
regulation depends on DNA-protein and protein-protein activation/repression
interactions affect conformation of DNA and change access to gene
control elets are combinatorial- mix and match monomers for control
173
how are SSB proteins regulated
```
alter TF-ligand binding conformation
regulate entry into nucleus/access to DNA
regulate amount of TF in cell
regulate DNA binding action of protein
phosphorylation/dephosphorylation of TF
```
174
how are nuclear hormone receptors controlled
Zn finger DNA binding motif;
steroid hormone enters cell and binds to nuclear hormone receptor, leading to conf change, recruitment of coactivators/repressors, and entry into nucleus
175
how does tamoxifen act in breast cancer therapy
nuclear hormone receptor activity:
acts as an antagonist to estrogen
doesn't allow dimerization, and prevents transcriptional effects from estrogen receptors
176
how is SSB protein regulated by nuclear entry
NF-kB is normally bound to IkB hiding the NLS, which holds it in the cytoplasm
if IkB is phosphorylated, it's targeted for degradation; degrading IkB shows the NLS to migrate into the nucleus and affect transcription
177
How does aspirin act on IkB?
it blocks IkB phosphorylation, blocking IkB degradation
| and it is anti-inflammative
178
how can amount of activators/repressors be regulated in a cell
and ACP gene as an example
specific genes can target the activators/repressors for degradation
ex. ACP gene targets beta-catenin via ubiquinin pathway and phosphorylation/degradation
(colon polyps caused by insufficient ACP activity)
179
how can DNA binding activity of a SSB protein be inhibited via Id proteins
Id proteins have helix-loop-helix domain, but no basic domain so can't bind to DNA
(HLH domain allows dimerization, basic allows DNA binding)
Id proteins heterodimerizing decrease efficiency of the helix loop helix proteins
180
what is a protein modification that can affect SSB proteins with CREB protein
ligand binds G-protein, phosphorylates CREB protein, recruits CREB Binding Protein (HAT), which recruts RNA Pol2 which leads to transcription
no phosphorylation= no CREB mediated transcription
181
4 mechanisms to control levels of gene expression
transcriptional regualtion
control of mRNA export from nucleus (cap and polyA tail)
control of mRNA degradation (siRNA action)
control of efficiency of translation (IRE/IRP at 5' UTR)
control of progein degradation (ie ubiquitination)
182
7 pieces of machinery that drive translation
```
ribosomes
mRNA
tRNA
aminoacyl tRNA synthetase
initiation factors
elongation factors
release factors
```
183
func of ribosomes in translation
site for mRNA translation into AAs
184
func of mRNA in translation
transcribed and spliced DNA copy that provides the 3 base codons that code for an AA's addition to a peptide chain
185
func of tRNA in translation
small piece of RNA with an AA on its 3' end
186
func of aminoacyl tRNA synthetase
enzyme responsible for adding the correct AA to the tRNA with the correct anticodon by ATP hydrolysis
187
func of initiation factors in translation
proteins needed to initiate ribosome assembly and mRNA translation
188
```
func of elongation factors in translation
ex. PTC and EF2
```
factos assoc with elongation ofa newly synthesized protein
ex. PTC catalyzes peptide bond via ATP coupled to tRNA
ex. EF2 uses GTP to move new peptide bond over in 3' direction one codon
189
func of release factors in translation
proteins taht bind to the stop codon can terminate polypeptide synthesis
190
start codon and what AA it encodes
AUG codes for methionine
191
missense, silent, and frameshift mutations in translation
missense- adds different AA
silent- different codon but same AA
frameshift- alters reading frame
192
4 phases of translation
initiation
elongation
termination
ribosome recycling
193
initiation of translation
2 subunits of ribosome brought together by a piece of mRNA
194
elongation of translation
codons are being read and corresponding tRNA is brought to A site of large ribosomal subunit
tRNA is then showed to P site and peptide bond is formed using PTC
new AA on the chain is moved via elongation factors and GTP to E site and uncharged tRNA is released
195
termination of translation
end of mRNA, release factors bind to stop codon and terminate protein synthesis
196
ribosome recycling of translation
ribosome is dismantled after termination to translate another mRNA
197
bac translation initiation
prokaryotes: use upstream shine-delgarno sequence to align mRNA start codon in P site of 30S subunit
IF1 and IF3 bind to 30S subunit
IF2 delivers formylmethionine to be 1st AA
GTP hydrolysis releases remaining initiation factors and binds 50S subunit
and next codon is placed in P site
198
eukaryote translation initiation
eukaryotes: Kozak sequences in intons that help find start codon
pre-initiation complex becomes initiation complex, then it scans down mRNA to find AUG
Initiation factors break off and large ribosomal subunit joins to form full ribosome
(5' UTR can affect initiation- stop or slow)
199
what is cap independent initiation
viruses can bypass the need for a cap to translate proteins
some viruses produce a protease that cleaves eIF to shut down cap-dependent translation
the virus can translate its genes by using the IRES (internal ribosome entry site) on RNA
200
what is interferon stimulation
when a virus invades a cell and the cell can't protect itself, it releases interferon into the extracellular space to come into contact w/ neighboring cells that recognize the interferon and translate anitviral proteins to protect themselves
201
what is mRNA editing
RNA code can be altered after it is made
| it can be translated differently in different parts of the body (one gene being truncated in certain areas)
202
what is rapamycin treatment
a drug that inhibits m-TOR protein translation
| it phosphorylates 4E-BP so initiation complex can't form
203
what is eIF2-alpha phosphorylation
when interferon is recognized by a cell, it phosphorylates eIF2-a to effectively turn it off
the enzyme can no longer bring in tRNA for translation
204
antibiotics and how they affect translation
streptomycin, tetracycline, erthromycin, chloramphenicol
| they inhibit translation by interfering with ribosome via tRNA binding, elongation, and peptidyl transferase
205
2 mechanisms to control iron regulation IRE and IRP
Iron response element IRE- RNA-stem loop structure in mRNAs that bind IRPs
Iron response binding proteins IRPs- bind iron and regulate expression of Ferritin and TFR
206
low iron vs high iron
low iron: little will bind to IRE-BP, so it binds to IRE. this stops Ferritin production and stabilizes transferrin receptor mRNA
High: iron will bind to IRE-BP and not to IRE, thus Ferritin mRNA is translated and TFR mRNA is degraded
207
function of ferritin and Transferrin receptor TFR
ferrin- sequesters iron (lowers free iron in cell)
TFR- transports iron into cell
low cell iron: up-regulate TFR translation and ferritin levels go down
208
func groups of an AA
alpha C with NH2, COOH, H, and R group
209
nonpolar/aliphatic R groups
MAGLIV
| not very reactive; made of hydrocarbons
210
aromatic R groups
FYW
211
polar uncharged groups
SCTPQN
| unbalanced electronegativities
212
polar charged groups
| positive and negative
positive: HRK (histidine, arginine, lysine)
basic with charged amines
negative: ED (glutamate, aspartamate)
acidic with resonant oxygens
213
func of disulfide bonds in proteins
S-S from 2 proximal cysteine residues stabilize tertiary and quaternary structure
ex. insulin, kertain in hair, and ribonuclease
214
post-translational covalent modification of AA sidechain
| hydroxylation of proline
in collagen, stabilizes structures,
mediated by Vitamin C
Scurvy is Vit C deficiency, therefore underhydroxylated collagens (weak)
215
post-translational covalent modification of AA sidechain
| carboxylation of glutamate
on prothrombin,
mediated by Vitamin K
required for effective blood clotting
Vit K deficiency leads to improper clotting
Warfarin prevents glutamate carboxylation,
therefore is an anticoagulant
216
post-translational covalent modification of AA sidechain
| glycosylation of asparagine
of proteins on cell membranes and that are secreted
increases hydrophilicity
congenital disorder of glycosylation (CDG) has malfunctions in this mech
217
post-translational covalent modification of AA sidechains
| acetylation and deacetylation
of histones in gene regulation (HAT and HDAC)
| cancer treatment can involve blocking HDACs
218
post-translational covalent modification of AA sidechain
| reversible phosphorylation
```
via kinases (adds P) and phosphatases (removes P)
affects signal transduction
```
Gleevec (tyrosine kinase inhibitor) is a cancer treatment- competitively inhibits bcr-abl kinase so substrate isn't phosphorylated so tumor cell can't proliferate via a abberant bcr-abl gene
219
post-translational covalent modification of AA sidechain
| ubiquitin
added to protein
signals them to be sent to the proteosome for destruction
multiple myeloma- velcade inhibits a proteosome that degrades good proteins
220
3 covalent bonds that make up backbone of polypeptide chain
peptide bond C1-N; partial double bond character; rigid bond
alphaC - peptide C; free rotation
amide nitrogen - alphaC; free rotation
221
2 models relating protein structure to function
AA sequence dictates structure-->function
1-lock and key- protein only fits specific substrate
2-induced fit- conformational change
222
mutations in AA sequence effects
some are polymorphic- differences in non-essential AAs
mutations can alter hydrogen and sulfur bonding --> vastly different structure and function
collagen and keratin need their strength
223
func of proteases
break down peptide bonds via hydrolysis
general: trypsin, chymotrypsin, pepsin (food digestion)
specific: help to activate precursor proteins to their active form (blood clotting factors are proteases that activate factors to yield clots after a trauma)
ex. angtiotensinogen cleaved by Renin to form Angiotensinogen 1, which is cleaved by ACE to Angiotensinogen 2, which is active hormone regulating bp
224
h bonds' role in secondary structure
weak, but numerous bonds between H and O
H bonds between amide N and carbonyl O are the driving force for secondary structure
alpha helix- H bonds every n and n+4
beta sheet- H between 2 chains
225
2 major secondary structure types
```
alpha helix- about 30% of all structures
ALL bio helices are right handed
all sidechians point outward
ex. hemoglobin
Thalassemia- disrupted alpha helices
```
beta sheet- about 30% of all structures
polypeptide chains held together by h bonds
anti-parallel are more common
ex. immunoglobulin or antibody
226
2 special protein structures
turns and loops
triple helix
turns and loops to compact proteins (small glycine and kinked proline)
triple helix in collagen- massive H bonding with lots of hydroxylated proline and small glycine
227
tertiary and quaternary structures
tertiary- spatial arrangement of polypeptide chain
globular (most, lipid or water soluble, diverse)
fibrous (long and either helices or sheets, insoluble, structure/protection)
N-terminus synthesized and folds first
proteins can assist in folding via chaperones
quaternary- multiple polypeptide chains to form 1 functional protein
ex. hemoglobin
228
role of loops in protein structure and func
enable polypeptide chain to form structures
can interact with other proteins with loops
mostly proline and glycine
229
how Kd represents binding strength
Kd is the dissociation constant
| Kd= CONC OF LIGAND when 50% of ligands are bound
230
how does heme enable myoglobin to bind oxygen
free or exposed Fe would be oxidized irreversibly, and the protein itself can't bind O2
a heme group is isolated w/in protein to work w/o producing free radicals
myoglobin is the main oxygen storage protein
231
carbon monoxide poisoning
CO has similar size and shape of O2
CO binds 200,000 x stronger than O2, so it outcompetes O2 binding
blocks myoglobin, hemoglobin, and mitochondrial cytochromes involved in oxidative phosphorylation
232
why is hemoglobin a good O2 transporter
myoglobin binds too strongly
hemoglobin has 4 heme groups interacting in positive cooperativity
Tense state: low affinity (in lungs)
high pH: high O2 binding (in lungs)
relaxed state: high affinity (leaving lungs)
low pH: low O2 binding (in tissues)
233
3 factors that cause protein denaturation
heat
pH
chemicals
234
Ribonuclease Refolding Experiment conclusion
protein was denatured, but regained full function
all info needed to fold protein is in primary AA sequence
cell environment not always necessary for folding
protein doesn't randomly explore all conformations (Levinthal's paradox)
235
2 classes of chaperones and their func
heat shock proteins (Hsp70)- induced at high T; binds to hydrophobic region to prevent aggregation, can help transport proteins across membranes in unfolded states
chaperonin- cap and two 7-subunit rings; binds to hydrophobic region of protein; uses ATP and conf change to partially fold protein so it can continue to final shape
236
why protein disulfide isomerase and protein prolyl isomerases are sometimes needed for protein folding
disulfide isomerase- enzyme needs to correct incorrectly bonded free Cysteines
protein prolyl isomerase- reforms proline from trans to cis for proper folding
237
4 diseases associated with protein misfolding
Alzheimer's
Prion disease
Parkinson's
amyloidosis
238
Alzheimer's disease w/ protein misfolding
normal AB-40 folds correctly
AB-42 misfolds and aggregates into amyloid plaques
leads to b-amyloid plaque and tau tangles
239
prion diesease w/ protein misfolding
prion protein misfolds, causing other prions to misfold
altered from alpha helices to beta sheets
causes aggregation within amyloid plaques
leads to neuron loss and gliosis
240
parkinson's disease w/ protein misfolding
beta-synuclein misfolds into Lewy Bodies
241
amyloidosis w/ protein misfolding
generalized protein misfolding in the rest of the body leading to variety of diseases
242
3 ways to purify a protein
size- gel filtration chromatography
charge- ion exchange chromatography
ligand binding properties- affinity chromatography
243
gel electrophoresis purpose and process
determines size of proteins
apply detergent- denature and uniformly coat proteins to be negative
use ladder, gel and electric field to separate
244
mass spec
determines sequence of unknown protein by molec mass
245
deman degradation
label and remove n-terminal AAs one at a time to identify
246
Western blots
use immunology to identify proteins on gel
transfer proteins to membrane, react with primary antibody, wash all unbound things, react with fluorescent secondary antibody to detect
ex. identify HIV infection (patient serum is primary antibody against HIV proteins)
247
palindromic double stranded DNA sequences
read the same forward and backward on compimentary strands 5' to 3'
likely to be cut by restriction endonucleases
248
southern blot
denature DNA then blotting the gel after electrophoresis onto a membrane that binds DNA
wash membrane with short label probe sequences of DNA or RNA to visualize
used when analyzing large quantity of DNA for particular sequences (diagnose genetic disease)
249
restriction fragment length polymorphism (RFLP) used to diagnose disease
RFLP: ex. looking for HbS mutation in sickle cell (in a restriction site)
digest patient DNA with diagnostic restriction enzymes
electrophorese against normal genome
southern blot with P32 labeled b-globin gene; look for 1 long band (abnormal) or 2 shorter bands (normal)
250
DNA fingerprinting for diagnoses
ex. paternal test
PCR with primers that surround variable number tandem repeat (VNTR) sequences
electrophoresis/detection of altered size of DNA fragment patterns
compare sample against target(s) for similarity match
251
transferring DNA, RNA, and proteins from electrophoresis gel to membrane
Southern blot- DNA
Northern- RNA
Western- protein
252
4 characteristics of hybridization probe that allow you to detect specific DNA sequences on a membrane
specific length- determines annealing T
specific sequence- binds to DNA of interest
something for visualization (radioactivity)
quantity- add enough to outcompete other strand of DNA in annealing to target strand
253
3 classes of enzymes used in recombinant technology
DNA polymerases- DNA-->DNA
reverse transcriptase- RNA--> DNA
DNA ligase- join DNA fragments
254
4 main steps repeated in PCR amplification
1- add termal stable DNA polymerase (Taq polymerase) and dNTPs and primers
2- denature DNA at 95 degrees
3- cool to 55 degrees for primer hybridization
4- warm to 72 degrees so polymerase copies the DNA
255
PCR use for diagnosis
prepare a primer that hybridizes with a mutant copy of DNA
carry out PCR
signal will amplify if genetic material has mutation
ex. cystic fibrosis, beta-thalassemia, etc
256
DNA sequencing
sequencing- use colored ddNTPs to stop synthesis and color last base; run through column to separate sizes; detector will read colors as they elute
257
PCR vs DNA sequencing purpose
PCR amplifies a DNA segment; uses double stranded DNA
DNA sequencing- determines sequence of unknown fragment; uses ssDNA, uses ddNTPs
both use primer sequences to initiation gene replication
258
6 cloning vectors
```
plasmids
bacteriophage
cosmids
BAC
YAC
retroviral vectors
```
259
plasmids-
vectors for amplifying DNA sequences in bac
| simple but inefficient
260
bacteriophage
vector used to infect E coli and use its replication machinery to produce the recombinant vector
more efficient than plasmid
261
cosmid
hybrid of plasmid and bacteriophage
| uses plasmid replication origin
262
BAC
bacterial artificial chromosome
| good for chromosome mapping and sequencing
263
YAC
yeast artificial chromosomes
| chromosome mapping and sequencing
264
retroviral vectors
can carry very large inserts
introduce DNA to mammalian cells
delivers gene therapy
265
use of mircoRNAs to measure mRNA levels
measure gene expression
they rely on hybridization, limited by background hybridization and signal saturation
can't provide sequence level info
can miss mutations, modifications, and splice forms
266
how do cells regulate size at the Restriction point
R point in G1 phase to check before S phase
somatic cells- make a decision whether hormones and GFs are present, and whether cell is large enough to undergo replication and mitosis
embryonic cells- R is bypassed; cells divide and get smaller
R point is 1st and most highly regulated checkpoint in cell cycle
267
main goal of somatic cell cycle
to ensure exact duplication of the genome in S phase followed by exact division of genomes in M phase to produce identical daughter cells
268
how do cells prevent re-replication of their genome in each phase
M- high CDK prevents building Pre-Replication Complex
G1- low CDK allows building of PRC
ORC proteins initiate replication by binding to the DNA binding origin and complexing with other proteins
S- high CDK activates replication and prevents PRC building
in short: pre-RC assembled during G1, activated during S, driven respectively by low and high CDK conc's in the cell
269
genetic instability causes in cell cycle
chromosome re-replication in S phase
or mis-segregation during mitosis
produces cancer or birth defects
270
end point of mitosis
2 identical diploid cells from 1 diploid cell
271
end point of meiosis
4 different haploid cells from 1 diploid cell
start: chromosomes A and a
1st- chromosome replication AA and aa
2- homologous recombination of chromosomes to provide genetic variation
3- separate homologs (meiosis 1: one cell AA and one cell aa)
4- separate chromosomes of daughter cells (diploid to haploid; 4 cells, A, A, a, a)
272
differentiated, post-mitotic cells stuck in the R point
continue to grow without cycling
| ex. neurons
273
CDK's role in cell cycle regulation
phosphorylate AA's
are regulated by cyclin
Active CDKs are produced by growth-factor hormones and result in cell replication and duplication
found at particular levels in particular cells (elevated in a tumor is indicative of proliferation)
low levels of RB or highly phosphorylated/inactive RBs indicate high replication rates (inhibits entrance into S)
274
CDI proteins
CDK inhibitors
turn off CDKs and repress cell cycle
mutations turn off the ability to turn off a CDK, leading to uncontrolled proliferation
275
mitogen
signal protein that leads to activation of CDK4 (leads to positve feedback to increase proliferation)
produces cyclinD
276
cell cycle checkpoint mechanisms
failing any checkpoint= cell cycle arrest
you can repair DNA at any phase (otherwise, apoptosis)
G1: R point; is cell big enough
S: should DNA be replicated?
G2: correct copies of new/old DNA?
M- did spindles form and act normally?
277
enzyme and rxn rates
molec that increase rxn rate without being irreversibly changed
specific structures and active sites
some use specific cofactors or coenzymes
classified and named by rxn they catalyze
278
Ea and reaction free E
Ea- E needed to overcome for reaction to proceed; dictates rxn rate
reaction free E- products - reactants E; dictates spontaneity
279
how enzymes work
enzyme binds and stabilizes the substrate in T state (induced fit changes conf pushing it toward product)
this lowers Ea needed to form product
280
3 types of active site chemistry to rearrange covalent bonds
metal ion chem- redox rxns
general acid-base catalysis- involves protons
covalent catalysis- transient covalent bond between enzyme and substrate
281
cofactor vs coenzyme
cofactor- metal ions required for enzyme activity
coenzyme- organic ligand that binds to the enzyme allowing it to act on substrate
(prosthetic group- tightly bound ligand;
holoenzyme- complex w/ the enzyme
apoenzyme- enzyme dissociated from complex)
282
Km vs Kcat
```
Km= CONC of substrate at which rxn is 1/2 vmax; use lineweaver-burke plot to determine
Kcat= turnover number; rate constant for # substrate molecs converted per time under sat condictions
```
Kcat/Km tells overall enzyme efficiency
large ratio indicates very efficient
283
4 types of enzyme inhibitors and func
competitive- inhibitor binds to substrate binding site
uncompetitive- inhibitor binds elsewhere; only binds to ES complex; lowers Vmax and changes Km
mixed/noncompetitive- inhibitor binds outside of active site to EITHER E or ES complex; affects both Km and Vmax
irreversible- permanent enzyme change (penicilin weakens bac cell walls)
284
4 types of enzyme regulation pathways
allosteric regulation- molec binds to enzyme and causes conf change to allow/disallow activity
covalent modification of enzyme- phosphorylation (via Kinases)
regulatory protein binding- enzymes bound by proteins to activate/inactivate
proteolytic activation- enzymes are inactive until cleaved by another enzyme (trypisinogen --> trypsin)
285
endogenous and exogenous dsDNA breaks
```
meiosis
generation of immune receptor diversity
DNA replication nicks
insertion of retroviruses into genomic DNA
ionizing radiation damage
medical tests
```
286
2 mechanisms of dsDNA break repar
NHEJ- can occur any time
doesn't restore orignal DNA (cuts some out)
used with immune receptor diversity
HR- S, G2, and meiosis
goal is perfect repair using sister chromatid
used with everything else
both: repair damage from ionizing radiation
287
NHEJ mechanism
Ku recognizes ds break
Ku recruits DNA-PKcs, recruits Artomis to form complex
nuclease to remove DNA if there's damage
polymerase fills gaps
repair finishes with ligase
regulated by 53BPI
288
HR mechanism
requires homologs
perfect repair- uses unbroken sister chromatid template to repair break
forms Holliday junction to repair breaks
junction is resolved to regenerate 2 ds DNAs
2 ways to cleave junc- results in cross over or non-crossover
regulated by BRCA1
in some cases of HR, use of other homolog as template (vs sister chromatid) can lead to LOSS OF HETEROZYGOSITY
289
how do defects in ds break repair lead to to cancer risk
mis-repair leads to genetic instability
| can lose signaling proteins (signals and transducers) and lose enzymes that mediate their repair
290
3 classes of proteins that repair ds breaks
```
sensing (Ku)
signal transudction (ATM)
DNA repair (HR)
```
291
next-generation DNA sequencing-
| short read sequencers
produce million-billion 100bp reads in a single run
low error rates (10-6)
commonly for high read-depth over specific subset of DNA template from complex mixture
292
next-gen DNA sequencing
| long read sequencers
produce about 10k reads of 10k+bps
high error rates (10-1)
useful in human genetics for linkking contiguous polymorphisms on same haplotype
single-molec approach to extract info from individual DNA template- "watches" single DNA polymerases
measures in close to real time- very long sequences can be generated
293
next-gen data collection
read depth
error rate
contiguity
read depth AKA coverage- number of times a genome is sequenced (higher= high confidence)
error rate- quality of converting signal to observation
contiguity- linking variations with e/o, usually across long distances
294
SNP identification criteria
ploidy need enough coverage to determine heterozygosity
295
exome sequencing experiments
sequence the 1% of genome that codes for proteins
| can identify novel coding SNPs among patients with same genetic disorder
296
which types of diseases are best characterized by exome sequencing
single-gene Mendelian diseases
variants are RARE, and not likely to be in existing database of variations
variants are non-synonymous
eg. Miller syndrome- embryonic exposure to methotrexate
297
prion structure normal vs infectious
normal- alpha helices
| infectious- beta sheets; hydrophobic; clump into lesions
298
how prion disease can be acquired
sporadic- random misfolding; infects other prions
inherited- familial strains; lead to early-onset of Creutzfeldt-Jakob Disease (fatal insomnia)
infectious- from diet or iatrogenetic (medical treatment)
299
bovine spongioform encephalopathy vs variant creutzfeldt-jakob disease
BSE is mad cow disease
vCJD- strain of prion that originated in cows and led to human vCJD via injestion
300
explain concept of prion strains
distinct characteristics- incubation time, clinical signs, distribution of protease resistant Prp in brains
all strains have same sequence as PrP
ex. hyper and drowsy strains tested in hamsters
301
pathophysiology of Alzheimer's
A-42 is incorrectly cleaved protein that incorrectly folds and becomes insoluble and leads to amyloid plaques
tau protein forms fibrillary tangles
tau and A plaques lead to inflammation and neuronal loss
occurs via post-translational amyloid precursor protein processing
302
possible Alzheimer's interventions
inhibit all APP products (which increase amyloid deposition)
use antibodies that attack A-42 (so it's never misfolded)
303
miRNAa
forms hairpin structure (double stranded)
don't allow translation
bind to mRNA in cytoplasm
processed in nucleus by drosha and dicer
perfect fit- RNA degradation
imperfect fit- degradation or translational repression
degradation- RISC complex with argonaut to act on RNA (endonuclease)
304
siRNA
from dsRNA cleaved by dicer, loads RISC complex- binds and cleaves; unwinding for recognition; cleavage
leads to:
chromatin remodeling (transcription silence)
regulation of transposons from viral infection
degradation
305
piRNA
comes from ssRNA (no dicer)
| regulates transposons with piwi and RISC
306
lnRNA
lots of functions
| regulates cardiac and skeletal muslces and immune response
307
2 nc regulatory RNA therapeutics
potential to target any RNA thus any protein
RNAi- target specificity; target proteins that can't be reached directly
antisense oligonucleotides ASOs- 4 types
focus on easy-access places (liver, eye)
308
exon-skipping ASOs
use oligonucleotides to force a truncated exon to be skipped so cell will join other normal exons and put back into reading frame
works with muscular dystrophy
309
anit-miRs and miRNA mimics
oligonucleotides used to antagonize or mimic miRNA
used to express something you don't want being repressed
works with Hep C virus targeting the liver
