lecture 1 Flashcards
1
Q
Definition of Recombination
A
- breaking and rejoining of DNA molecules in new combinations
- rapid evolution
2
Q
State size and minimum region of oriC
A
- 245 aa
- 2/3 of oric - 4/5 of a 9 bp seq
- 1/3 of oric - of 3 copies of AT rich 13 bp seq
- 14 copies of GATc
3
Q
Mechanism of SOS response
A
- lec A bind s to lex A boxes and repress SOS operons
- damage activates Rec A to form Rec A star
- / rec A interact with lec A and cleave lec A / repair the damage
- become Rec A again and stop autocleaving
4
Q
Haloenzyme and core enzyme
A
alpha 2 beta beta prime omega sigma ithout sigma unit
5
Q
State the role of sigma factor in RNA polymerase
A
- make it binds tightly to promoter and melt the DNA( allow RNA polymerase to start copy)
6
Q
State the subunits of RNA polymerase
A
alpha, beta, beta prime, sigma, omega
7
Q
Chromatin immunoprecipitation
A
- crosslink DNA binding proteins with formaldehyde
- shear DNA along with the bound proteins into fragments
- add antibodies which binds to the DNA-binding protein, to isolate the complex by precipitation
- amplify the precipitated DNA with PCR
8
Q
Ways to identify Transcription factor binding site
A
-Chip and next generation sequencing
know gene sequences of both sides of TBS but not at TBS
9
Q
Microarray analysis
A
- label mRNA with fluorescence dye
- hybridize with cDNA
- if there is mRNA, binds with cDNA(probe)
- analyse the gene expression
10
Q
State process of RNA sequencing using next generation sequencing
A
- RNA to cDNA
- add adaptors to cDNA
- measure gene expression by counting RNAs from each exons
- allow the identification of gene splicing variants
11
Q
State the role of microarray analysis
A
- analyse expression of thousand of genes at the same time
- can pattern of gene expression at different times different tissues or different conditions
12
Q
State definition of regulons
A
- combination of many operons and single regulated gene which is controlling
13
Q
Global regulation
A
-control system which regulate many genes globally in response to single environmental factor
14
Q
Catabolite repression
A
-glucose suppress less preferred carbon sources
15
Q
Lac A
A
- beta-glycosidase trans-acetylase - detoxify beta glycosides by acetylation
16
Q
Position where proteins bind to DNA
A
CAMP-CAP footprint - (-72 to -52)
RNA polymerase footprint: (-50 to +17)
Repressor footprint: -5 to +21
17
Q
Euchromatin , heterochromatin
A
Euchromatin - less condensed
Heterochromatin - condensed
18
Q
State the components of nucleosome
A
H2A, H2B, H3 and H4
19
Q
State how histone structure forms
A
- H3 and H4 form tetramer
- (x2) H2A and H2B dimers
- from histone octamer
20
Q
Windograsky column
A
- wide diversity of ecosystem in test tube ( soil, sediment , water, carbon)
- contains aerobes and anaerobes
- carbon fixation and fermentation
21
Q
Structure of nucleosome AND size
A
- histone core and inverted repeat DNA
- 10nm
22
Q
Three types of functional elements require for maintaining chromosome replication and stability
A
- telomere
- ORI
- centromere
23
Q
Reassociation kinetics
A
- to predict the number of genes per genome
24
Q
State the composition of human genome
A
- repeated sequences ( single sequence repeats, segmental duplications, transposons), unique sequences( genes, introns, non- repetitive DNA(neither introns/codons))
25
snRP
small nuclear ribonucleoprotein particles- small nuclear RNA with at least seven protein subunits
26
how is splicing specific?
- specific nucleotide sequences
27
Experimental evidence of splicing / splicing products
- unspliced pre mRNA ( 497)
- lariant plus exon - 339
degradation product - 252
- correctly spliced mRNA - 367
- only lariat intron : 130(non linear)
28
State the proteins required for slicing other than spliceosome
SR proteins bind to exonic splicing enhancer(ESE) - mark exon regions
hnRNP - marks intron regions
29
hnRNP
heterogenous nuclear ribonucleo protein
30
Two types of elongation factor and their size
Ef-Tu(45kDa) and EF-T(30)
31
What does the probability of forming a protein with no errors depends on?
the number of amino acid (n) and the frequency of insertion of a wrong amino acid (p= (1-E) to the power of n
32
Which molecular chaperones are needed to favour correct folding and how do they do it?
- Hsp40 and then Hsp70
- hydrophobic patches recognised by Hsp40( heat shock protein 40 family members)
- deliver the substrate to ATP-bound(open) conformation heat shock cognate protein 70
- causes ATPase activity of Hsc70 which shield the hydrophobic patches of the substrate, allowing time for them to fold properly
- nucleotide exchange by nucleotide exchange factor and release of polypeptide
33
State the multiple possible fates of Hsc70
- released and find its stable conformation
- passes on to other chaperones for further folding or form multimeric complexes
- degraded by proteosomes ( non-productive)
- transporting to other organelles
34
Hsc70 co-chaperones
Hsp40 family members, CHIP
35
structure of chaperonin and function ( like two circles stack)
- 14x Hsc60
| - isolate small protein from cytosol
36
How is the protein fate determined?
- interacting, competing network of co-chaperones which determine the fate of a chaperone client but not pre-determined
37
Roles of cytosolic molecular co-chaperones
- prevent aggregation of unfolded proteins
( allow time for them to fold and shields hydrophobic regions)
- allows them to fold further or form multimeric complexes (assembly of histone complexes)
- provide controlled environment for proteins to fold ( chaperonins isolate proteins from cytosol to be able to fold properly)
- direct unfolded proteins to proteosome for degradation( Bag-1)
38
ADP ribosylation and one example of ADP-ribosylase
- adding ADP ribose residues to protein
| - Diptheria toxin is an ADP-riboylase that targets EF-2
39
Secretory system modifications
- O-glycosylation
- N glycolsylation
- Proteolytic cleavage
40
Why are N-glycans attached to some ER proteins?
- increase solubility due to many hydrophilic residues
- influence folding rates and final protein conformation/influence the activity of protein or its interaction with other molecules ( due to constrain the alpha-carbon backbone of the polypeptide
- act as flags to interact with ER chaperons and communicate on the stage of folding.( Remove one glucose-still folding, remove all - move out of folding stage)
41
How complex are the leader peptides?
- two targeting signal
| - one into mitochondria, on into intra-organelle compartment
42
Example of complexity of LP - to enter metro , OM and IM
- all have matrix-targeting sequence of LP to enter outer membrane
- cleavage by matrix protease and IMS protease to get to inner membrane
- cleavage by matrix protease only to get into matrix
43
Dual targeting
- weak SP or LP which can target to ER by binding to SRP( co-translaationally) or to mitochondria ( post-translationally)
44
translational control mechanisms
1. Regulation of the activities of initiation and/or elongation factors by
phosphorylation (pro- and eukaryotes).
2. Blocking/opening of ribosome binding sites by reversible changes in
secondary structure (prokaryotes).
3. Autogenous regulation. Protein product of a gene binds to ribosome
binding site in mRNA, preventing initiation on (prokaryotes).
4. Reversible binding of a repressor protein to a response element in 5’
UTR (eukaryotes).
5. Differential stability of mRNA
