Topic 4 Flashcards
1
Q
DNA is ….
A
inert
2
Q
RNA is the….
A
heart of transcription
3
Q
copied strand
A
coding strand
4
Q
other strand
A
template strand
5
Q
RNA builds
A
5’ to 3’
6
Q
Which base is replaced in RNA and with what
A
T replaced with U
7
Q
RNA polymerase core enzyme made up of …..
A
5 subunits: 2 alpha, 2 beta, w
8
Q
RNA holoenzyme includes….
A
additional sigma unit
9
Q
What does core enzyme do?
A
unwinds DNA and forms transcription bubble
10
Q
What stops the RNA
A
transcriptional terminator
11
Q
Intrinsic (rho-independent terminators)
A
Forms when RNA hairpin structures form, creating a U that pause RNAP then forces it off
12
Q
RNAP
A
RNA polymerase
13
Q
Rho-dependent terminators
A
protein called Rho that cause RNAP to dissociate after certain sequence
14
Q
What initiates transcription
A
promoters
15
Q
How do you is a promotor is activated
A
binding of sigma factors and regulatory proteins
16
Q
housekeeping sigma factor
A
sigma70 or RpoD
17
Q
Rpod
A
recognizes 2 sequences upstream of transcriptional start site, -35 and the Pibrew box
18
Q
transcriptional start site
A
+1
19
Q
Pribrew Box
A
~10 bp upstream of +1 site
20
Q
Messenger RNA
A
converted to protein via translation
21
Q
Transfer RNA
A
functional RNA used in translation
22
Q
Ribosomal RNA
A
functional RNA used in translation
23
Q
open reading frames (ORF)
A
sequences translated into proteins
24
Q
untranslated regions (UTR)
A
parts of mRNA not translated
25
Polycistronic
encodes more than 1 polypeptide
26
Operons
cluster of genes with 1 promoter
27
Genes in an operon are.....
cotranscribed
28
5' UTR
+1 to start codon of gene
29
ORF
start codon to stop codon
30
3' UTR
stop codon to final transcribed residue, includes termination sequences
31
how is transcription different in eukaryotes (8)
- 3 polymerase
- more complex, 12+ subunits
- requires transcription factors to bind DNA and recruit RNAP
- do not use operons, each gene has own promoter
- mRNA spliced to remove introns
- 3' polyadenylated and G cap at 5'
- mRNA transcribed in nucleus, translated in cytoplasm
- TATA box
32
Archea Transcription
| less complex version of eukaryotic transcription
- resembles RNA pol II
- uses TATA box
- uses operons
33
Protein Structure?
- made of amino acids connected by peptide bonds (polypeptides)
- run N----> C
- only get built at the C end
34
Name the 2 Rare Amino Acids
1. Selenocysteine
| 2. Pyrrolysine
35
How are amino acids groups? Name the four groupings!
- grouped based on R-groups!
1. non-polar (hydrophobic)
2. polar
3. positively charged
4. negatively charged
36
Primary structure of proteins?
chain of amino acids
37
Secondary structure of proteins
alpha helices and beta sheets
| - formed by H bonding of peptide backbone
38
Tertiary structure of proteins
3D structure of protein
| - many secondary structures arranged together
39
Quaternary structure of proteins
many polypeptides coming together
| ===> called multimeric proteins
40
what are subunits in proteins?
the individual peptide chains in multimeric proteins
Identical units: homomeric
Different: heteromeric
41
What are protein domains?
- structural/functional segments in proteins
| - can be small/large
42
Example of a protein domain?
Helix-turn-Helix (HTH) domains
- bind DNA (DNA binding regulatory proteins)
- found in many proteins of any Salmonella genome
43
what does tRNA do?
- convert/translate the mRNA sequence into a protein sequence
44
Each tRNA has a specific ________ that binds a particular three-base codon.
anticodon
45
What are tRNA synthetases?
- enzymes that 'charge' tRNAs (add amino acid to the CCA at the 3' end)
46
How many base pairs in a codon?
3
| keep going guys, we got this!!
47
T/F: The same tRNA can be used for two different codons.
True!
| - some tRNAs work using the 'wobble' method by having a mismatch in the 3rd position
48
the typical Start codon?
typically: AUG
| ATG in DNA sequence
49
alternative start codons?
GUG, UUG
| these aren't labelled on the codon table as start codons so we memorize them?? Uhh
50
What is the start codon translated to in bacteria?
N-formylmethionine (fMet) using a special tRNA
51
What are the ribosome subunits?
| in prok.
large: 50S, small: 30S ===> total is the 70S ribosome
52
What mainly carries out the functions of the ribosome?
rRNA! It also catalyzes peptide bond formation
53
What are Shine - Dalgarno sequences?
the ribosome binding sites for translation initiation
54
What is used for energy in translation?
GTP!!!!
55
How does translation begin? (initiation)
- the ribosome binding site (shine dalgarno) binds to 16S rRNA in a random 30S ribosomal unit
- this helps robosome look for AUG codons (bona fide step start)
- fMet tRNA binds + GTP used for energy ===> 50S subunit recruited
56
```
Translation Initiation
(dumbed down version)
```
1. Shine Dalgarno finds random 30S small subunit for bon fide (which is just looking for AUG codon)
2. fMet tRNA binds AUG and its ready to hunt down a 50S subunit
3. 50S subunit found and bound --> ribosome is ready to translate!
57
The three sites of the ribosome
EPA
A site: new tRNA enters
P site: where the growing peptide chain is
e: exittttt! uncharged tRNAs leave
58
What is the order of steps in translation?
1. Initiation
2. Elongation
2. Termination
59
Translation Termination
- ribosome encounters stop codon so a release factor binds and dissociates ribosome
60
What are polysomes?
Many ribosomes on one transcript.
61
Can RNA be translated while transcription is still ongoing?
Yup. This is coupled transcription/translation in prok.
62
T/F Euk. ribosomes are smaller than prok. ribosomes.
False! They're bigger --> 40S small, 60S large
63
Differences between euk. and prok. translation
- in euk. transcription/translation is NOT coupled
- euk. mRNA only encodes one gene
- 5' cap recognized in euk.
- euk. translation is slightly more complex/more regulation
64
Chaperones
| hint: 'changing rooms'
- help proteins to fold properly
- have controlled environment for proteins to fold properly
- use ATP hydrolysis!!!
65
E. coli chaperones to know
DnaJ/DnaK, GroEL and GroES
| - most abundant chaperones in E.coli cells
66
Where are proteins synthesized in prokaryotes?
in the cytoplasm by ribosomes
67
How are proteins transported outside the cell if they're required elsewhere?
reminder: proteins are impermeable to membrane
translocase systems! they help move proteins across and into cytoplasmic membrane
68
Two core secretion systems?
| hint: sec tat
1. Sec secretion system
| 2. twin arginine translocase (tat)
69
Sec secretion system
secretes UNFOLDED proteins and then the protein folds once its at the place it needs to be
70
Tat pathway
secretes FOLDED proteins
71
Regulating Transcription Initiation?
control whether or not RNA polymerase binds a
| promoter and initiates transcription
72
Transcription Factors?
- DNA binding regulatory factors
| - dictate if RNA polymerase is able to regulate transcription at a given site
73
Purposes of DNA binding domains?
- dimerization
- interacting with other proteins (RNA pol)
- regulatory domain
74
Activator
Transcription factors that promote transcription
75
Repressor
transcription factors that inhibit transcription
76
________ work by binding
DNA at promoter & recruiting
RNA polymerase (sigma factor)
to begin transcription
Activators
77
__________ bind DNA &
prevent RNAP DNA binding or
transcriptional initiation after it
binds.
Repressor
78
What does it mean to be regulated allosterically?
binding of an
effector activates or
inactivates protein
79
Inducers
turn on activator proteins or inactivate repressors
80
Co-repressors
activate repressor proteins
81
Inducible System?
is one that is off by default, but
| can be turned on
82
Repressible System?
one that is on by default, but
| can be turned off
83
What is ArgR?
repressor protein that controls the expression of an arginine
biosynthesis operon
84
Explain Arginine Biosynthesis?
Low arginine levels = ArgR isn’t bound by arginine, doesn’t bind DNA –> arginine is synthesized
High arginine levels = ArgR to bind
the Operator & prevent
transcription of this operon
85
Lac operon?
Machinery for breaking down lactose
86
LacI repressor protein
binds lac Operator prevents transcription
87
Catabolite repression
microbes eat best nutrients first, shut off secondary nutrients until needed --> not wasting energy
88
T/F: In the presence of glucose,
production of cAMP is
inhibited.
True!
89
What is required for lac operon to be expressed? (2 important things)
lactose AND low glucose levels
90
cAMP?
signalling molecule or second messenger
91
Stringent Response
ppGpp (produced in response to amino acid starvation) shuts down protein synthesis & induces amino acid biosynthesis
92
Quorum sensing?
sensing the local density of cells through secreting/detecting specific molecules
93
Autoinducers?
Quorum sensing involves signalling molecules (form of chemical communication)
94
Signal Transduction uses which two proteins:
Sensor Kinase and Response regulator
95
Sensor Kinase?
-resides in cytoplasmic membrane. -Senses specific signal -> activates kinase activity -> adds phosphate to response regulator
96
Response Regulator?
- When phosphorylated, becomes active
| - Binds DNA to regulate expression of target genes (activator and/or repressor
97
Transcriptional Silencing?
very tightly shutting off
| expression of genes by altering the genome structure at promoter regions
98
Example of transcriptional silencer:
H-NS (E. coli, Salmonella,
| other Gram-negative bacteria)
99
Role of H-NS:
prevents binding & restructures
DNA to a rigid structure to prevent RNA
polymerase from binding the DNA
100
H-NS binds to ...
regions of genome with high AT%
101
Counter-silencing?
DNA-binding activators bind specific silenced loci & reverse effects of H-NS (re-structure DNA and/or remove H-NS)
allow specific genes to be expressed
102
Global Regulators?
regulate large numbers of different genes in response to a given signal or environmental cue
103
Regulon?
complete set of genes controlled by a given regulator
104
Transcriptional attenuation
regulation that involves prematurely
| terminating mRNA synthesis
105
Translation efficiency?
usually whether or not
| RBS is free to be bound by the ribosome
106
Two major RNA structures that “shut off ”
| gene expression
- Formation of a stem-loop structure involving RBS
| - transcriptional terminator
106
Two major RNA structures that “shut off ”
| gene expression
- Formation of a stem-loop structure involving RBS
| - transcriptional terminator
107
Ribonuclease (RNase)
degrade mRNA
108
sRNA?
- base-pair to target mRNA(s) to regulate half-life and/or translational initiation
- affects RBS availability and/or RNase targeting
109
Hfq?
RNA chaperone that binds to both RNAs to stabilize their interaction
110
Riboswitches?
- ligand-binding RNAs –> adopt intricate 3D structures that specifically bind a particular small molecule
- allows riboswitches to sense their environment
111
Mechanism of Riboswitches?
Aptamer binds ligand, changes base-pairing in
the 5’UTR of mRNA to affect formation of:
(i) A stem-loop that
sequesters the RBS (blocks translation)
(ii) a transcriptional terminator
that prevents transcription of the genes.
112
Feedback inhibition?
An enzyme within a given biosynthetic pathway is sometimes inhibited by
the end product of that pathway
113
Protein-Protein interaction?
one protein binds another to control its activity
114
Post-translational modification
enzyme adds a chemical moiety to a specific amino acid residue of a
protein, which alters its activity (change activity, or turn it “on”/“off ”)
115
Role of Proteases?
enzyme used for clearing away and recycling misfolded proteins
