Exam 3 Flashcards
Glutamine synthetase
AMP added and activity decreases
Peptidyltransferase
23S rRNA
Catabolite repression
If glucose and lactose are present in the environment
Phosphatase
Mediates feedback for Two Component Regulatory Systems
Always removes phosphate group at constant rate
Riboswitches
A metabolite bonds mRNA on 5’ end, hiding the Shine Dalgarno sequence, preventing translation.
EF-G
GTP used to translocate the ribosome along the strand
Catabolite Activator Protein
Bonds to CAP binding site
Only occurs when bound by cyclic AMP
Mutation frequency
Relatively rare
Every 10^9- 10^8 base pairs
DNA Polymerase has proofreading capabilities
Termination codons
3 codons
tRNA will not bind but a release factor will bind and cleave the peptide. Ribosomal subunits dissociate.
Antisense RNA
40-490 bases long (avg 100)
Has its own gene and can bind to multiple different mRNA’s
Shuts down translation by targeting strand for deletion
Cyclic AMP
Synthesized by adenylate Cyclase
Adenylate Cyclase
Synthesizes cyclic AMP
Why are mutations more common in Prok than Euk?
Less DNA
HAPLOID, so the DNA will be passed on indefinitely.
Release factors
Euk=> 1
Bact=> 3
Hfq protein
Facilitates proper RNA to RNA interaction in terms of antisense RNA
Mutant strain
Genotype will differ
Phenotype may or may not differ
Efficiently of translation
More efficient in prokaryotes because there is no nucleus, so there is no separation between transcription and translation
Multiple ribosomes can bind to one mRNA
Regulatory proteins
Found in major groove
Homodimeric
Structural motifs help bind to DNA:
Helix-turn-helix, Zinc finger, Leucine zipper
DNA Polymerase III in Arch. and Euk.
B=> replication
A&C=> repair
DNA Polymerase III in Prok.
A&B=> repair
C=> replication
DNA Polymerase III
Needs a primer
Goes 5’ to 3’
Proofreading capabilities
3’ to 5’ exonuclease activity
topoisomerase
Relieves supercoiling
primase
Lays down an RNA primer
Getting ready for DNA Polymerase at
Origin of Replication
DnaA binds
DnaB (helicase) binds: pulls apart double helix
Single-stranded binding proteins
Okazaki Fragments in Prok.
1000-2000 bases
Okazaki Fragments in Euk. and Arch.
100-200 bases
DNA Polymerase I
Removes RNA primers
Has 5’ to 3’ exonuclease activity
Add DNA
DNA Ligase
Connects DNA fragments together
Theta structure
What allows bacteria to divide so rapidly
DNA replication can start in many “layers”
Termination Site (Ter Site)
Tus protein binds and physically disrupts replication forks
Topoisomerase also involved
Tus protein
binds and disrupts replication forks
Arch. # of Origin of Replications
multiple
Prok. # of Origin of Replications
one
RNA differences to DNA
Contains Uracil instead of Thymine
Single stranded
Has ribose
Transcription
RNA polymerase
Promoters help RNA to bind
RNA Polymerase
5 subunits
σ factor
Recognizes the promoters exposed in major grooves
Leaves after recognition
Also involved in: endospore formation, different consensus sequences
Promoters
Always upstream of gene start site
Pribnow Box and -35 Sequence
Pribnow Box
Upstream 10 base pairs
-35 Sequence
TTGACA
Upstream 35 base pairs
TATA Box
Upstream 30 base pairs
Only Arch and Euk
RNA Polymerases in Arch.
Only one
Similar to Euk’s II
RNA Polymerases in Prok.
Only one
Not similar to any others
RNA Polymerases in Euk.
I, II, and III
II similar to Arch.
Intrinsic terminators
Based on mRNA structure
Hair-pin shaped loop
Inverted repeats
Hair-pin shaped loop
Immediately following gene is a stretch of U
Before that are sequences of a.a. in opposite order
Interacts with RNA polymerase and breaks
Rho-dependent transcription termination
RUT Site=> Rho Utilization Site
RNA polymerase pauses when it reaches loop
Rho protein then cleaves the strand
mRNA in Euk.
monocistronic
mRNA in Prok and Arch
polycistronic
Introns in Euk.
Do not lead to specific proteins
Introns in Prok.
NO INTRONS
Exons in Euk.
Lead to specific proteins
Ribonucleoproteins
snRNP’s
Remove introns in Euk.
5’ Cap in Euk.
7-methyulguanosine cap
Initiates translation procedure
Poly A tail in Euk.
At 3’ end
200 bases long
Poly A tail in Prok.
Means it’s targeted for degradation
10-40 bases long
How many different codons?
64
Wobble Site
Third spot in the codon
Shine Dalgarno Sequence
16S rRNA finds this sequence at 5’ end before start codon and it complimentary base pairs with a.a. on 3’ end of 16S rRNA
Amino-acyl tRNA Synthetase
Adds in amino acid
Located on 3’ end of tRNA
tRNA
90 nucleotides long
folds make it more stable
Large subunit of rRNA in Prok
50S
5S and 23S rRNA
Small subunit of rRNA in Prok
30S
16S rRNA
Prok. Ribosome
70S
Euk. Ribosome
80S
Large subunit of rRNA in Euk
60S
28S, 5.8S, and 5S rRNA
Small subunit of rRNA in Euk
18S rRNA
Formyl-Methionine
AUG Initiator Group
Attached first in prok
Initiation of Protein Synthesis
30S subunit binds
Formyl-Methionine attached
Then, 50S subunit binds
DNA designation for proteins Euk
3% of DNA
DNA designation for proteins Yeast
70% of DNA
DNA designation for proteins Bacteria
90% of DNA
A T
2 H-bonds
C G
3 H-bonds
DNA A Form
11 bases per turn
DNA B Form
10 bases per turn
Histones
Euk and Arch
Supercoiling comes form wrapping around these
DNA Gyrase
Negative supercoiling
Many antibiotics target this
Reverse DNA Gyrase
Positive supercoiling in hyperthermophiles
Adenylate cyclase presence of glucose
adenylate cyclase synthesis inhibited
cAMP transported out of the cell
Missense mutation
1st or 2nd place in codon changed
DnaK and DnaJ
ATP dependent
chaperones that allow for properly folded proteins
Negative control of transcription
Occurs when the DNA binding protein (repressor) inhibits initiation of transcription
Operator DOWNSTREAM
Repression or Induction
Adenylate cyclase absence of glucose
active
Nonsense mutation
stop codon created
usually results in an incomplete protein
GroEL and GroES
ATP dependent
for more stubborn proteins
only about 100 genes in E. coli need this
Operator
Where the repressor protein binds
Diauxic growth
Two phases of growth because changing of glucose to lactose
lag phase in between
Silent mutation
3rd or Wobble site changed
results in a normal protein
Constitutive proteins
always expressed
“housekeeping genes”
Repression
Anabolic pathway
Attenuation
Occurs DURING transcription
Only in Prok.
Feedback inhibition
Post-translational control
typically targets first enzyme
Induction
Catabolic pathway
Tryptophan plentiful
The ribosome will continue translation
Subunits 3 and 4 of mRNA will bind
RNA polymerase terminates and genes aren’t transcribed
Allosteric site
Once bound, the conformational changes prevents the binding of substrate
Arginine operon
REPRESSION (NEGATIVE CONTROL)
Excess Arginine acts as a corepressor and binds to the repressor to block RNA polymerase which blocks transcription
Tryptophan scarce
The ribosome will stall
Subunits 2 and 3 of mRNA will bind
RNA polymerase continues to transcribe genes
EF-Tu
Helps load tRNA’s into the A site
Active when bound to GTP
Concerted feedback inhibition
isoenzymes
different enzymes that catalyze the same reaction but are under different regulatory control
The Lac Operon
INDUCTION (NEGATIVE CONTROL)
Repressor binds when lactose is absent. When lactose is present, it acts as an inducer and binds repressor thus allowing transcription to proceed
Sensor kinase protein
Found in cytoplasmic membrane
typically autophosphorylates at the histidine residue
Ex) EnvZ
EF-Ts
Catalyzes the binding of GTP
Reactivates EF-Tu
Covalent modification of enzymes
AMP used to modify enzyme
Can also add ADP, PO4, CH3
Difference between Positive and Negative Transcriptional control
The binding location
Positive=> UPSTREAM
Negative=> DOWNSTREAM
Response regulator protein
phosphate transferred to response regulatory protein
cytoplasmic DNA binding protein
Ex) OmpR
Base analog
Is mistakenly integrated into the genome
Example of a chemical mutating DNA
ethidium bromide
Rec A protein
SOS Response
binds and holds gaps together
Sfi A Protein
SOS Response
inhibits cell division
DNA Polymerase IV and V
Used in SOS Response
carcinogenic
causes mutations
Bruce Ames
developed Ames Test in 1970’s
Mutational reversion assay
Salmonella typhimurium used (Histidine auxotrophs)
cell wall altered so it’s more penetrable by chemicals
repair mechanisms removed
Ames Test
Control Plate: Bacteria, molten agar with small amount of Histidine
Test Plate: Bacteria molten agar with small amount of Histidine, chemical to test mutagenicity
Auxotroph
only grows on a medium that provides a lacking amino acid or nucleotide
Morphological mutations
changes that can be seen
Lethal mutations
cannot survive
Conditional mutations
only seen under certain environmental conditions
Biochemical mutations
a biosynthetic pathway inactivated to change the biochemistry of the cell
interrupt an amino acid or nucleotide
Frameshift mutation
occurs when base pairs are deleted or inserted
alters the reading frame
Reversion mutation
when an earlier mutation is reversed by a second mutation
Same-site (true) reversion
converts the mutant nucleotide sequence back to the original sequence
Second-site reversion
a second mutation occurs at a different site in the DNA and causes the wild type phenotype to be restored
Induced mutations
results from exposure to known mutagens which are primarily physical or chemical agents that interact with DNA in a disruptive manner
Spontaneous mutations
a random change in the DNA arising from errors in replication
Ethidium bromide
an example of an induced mutation producing chemical
Ionizing radiation
radiation ejects orbital electrons from an atom and causes ions to form DNA BREAKAGES
Gamma rays (most penetrating)
Cathode rays (least penetrating)
Nonionizing radiation
Radiation that excites an atom to a higher energy state
UV radiation
absorbed by DNA and creates pyrmidine (CT) abnormal bonds in DNA
Ultraviolet light
only targets the surface
Nucleotide Excision
Involves UVR-ABC enzyme
removes distortion, usually 12 nucleotides
DNA Polymerase I fills in gaps
DNA Ligase fuses them back together
Excision Repair
Corrections to distortions in DNA
SOS Response
Works to repair problems in the absence of a template strand
ERROR PRONE LAST DITCH EFFORT
Base Excision
DNA Glycolsylase severs bonds between the two bases
AP endonucleases nick DNA
DNA Polymerase I removes and replaces base
Direct Repair Mechanism
Photoactivation
thymine dimers caused by non-ionizing radiation removed with photolyase
Photolyase
removes Thymine dimers caused by non-ionizing radiation
DNA Glycolsylase
severs bond between two bases during Base Excision
AP endonucleases
nick DNA during Base Excision
