What is required for the protein surface to bind to the DNA surface?
For the protein surface to be complimentary to the DNA surface
For the protein surface to be positive (DNA backbone's are negative)
For the protein surface to contain hydrophobic residues
In which groove can the order of base pairs be differentiated?
Major groove
Minor groove
The major groove is 15A wide
Which is smaller?
Proteins that bind in the major groove must be more specific than those that bind in the minor groove
Why is Arg such a useful residue?
It has 2 H acceptors, so it can bind to guanine
It has 2 H donors, so it can bind to guanine
It has 2 H acceptors, so it can bind to cytosine
Which of the following can bind to both G & A
Glu
Val
Asn
Tyr
What is base stacking?
Where hydrophobic bases stack to avoid water
Where hydrophobic bases pull the DNA inwards to avoid water
Where hydrophobic bases twist the DNA to avoid water
What is required for a protein to take advantage of base stacking?
Residues with delocalised electrons
Hydrophobic residues
Hydrphillic residues
For it to make a gap between bases
10bp covers a distance of 34 A
Describe DNA
Long, repetitious and thin
Long, non-repetitious and thick
Short, repetitious and thin
Long, repetitious and thick
Short, non-repetitious and thin
DNA can bend 180 dgrees over how long a stretch?
40
30
20
What does Phage Lamda Repressor bind to?
Phage DNA
Bacterial DNA
What is being shown here?
A half site
A palindromic sequence
A restriction site
How does Phage Lambda Repressor Work?
Binds to bacterial DNA at the lambda cro gene
Binds to phage DNA at the Lambda cro gene
And blocking RNA polymerase binding
Phage Lambda Repressor binds to only one target sequence
The Phage Lambda repressor binds as a _
Monomer
Dimer
Tetramer
Each subunit attaches to the DNA using a _ and binds to the other subunits using a _
Alpha helix, alpha helix
Alpha helix, nothing- it's a monomer
Beta sheet, beta sheet
Beta sheet, nothing- it's a monomer
Phage Lambda Repressor has multiple subunits because ...
It binds to a sequence long enough that the protein cannot be removed
It binds to a sequence long enough that the sequence becomes unique int the DNA
It binds more exactly - so it only has a single target sequence
Where does Phage Lambda Suppressor bind?
2 adjacent major grooves
2 adjactent minor grooves
An adjactent major & minor groove
32A apart
34A apart
36A apart
A non-specific interaction doesn't involve bonding with any particular base
List some specific interactions between Phage Lambda Repressor and DNA
Ser - G, and Gly - T,
Gly- T and Ser-G
Gly- C- and Ser-T
The DNA sequence is all read from the same DNA strand
A helix-turn-helix motif is often used for reading DNA
Different alpha helices (of the helix-turn-helix) motif perform specific and non-specific interactions
Proteins don't have to land on the DNA in exactly the right spot, they only need to...
Bind to the phosphate backbone
Bind to the bases
And then roll freely along the DNA
Methionine repressor...
prevents methionine production
prevents binding of methionine to tRNA
prevents methionine being added to the amino acid chain
The Methionine Repressor binding site is...
Palindromic
Almost palindromic
8bp
16bp
The binding site of most helix-turn-helix proteins are 16-18bp long
How does the Methionine repressor bind to the DNA?
Uses 2 alpha helices
Uses 2 beta sheets
One from each monomer
One from each end of the monomer
Into one major groove
Into two major grooves
Two beta strands have the same diameter as an alpha helix
Specific interactions in the methionine repressor happen between Lysine, Threonine, A & G
Restrictions enzymes are part of the bacterial immunity... how?
Recognises and destroys non-self DNA
Cuts around viral DNA that has infected the genome
Proteases break down the viral coat
Restriction enzymes and transferases come in pairs, with the restriction enzyme made first
How does EcoRI bind?
Phosphate backbone reactions in the minor groove
Phosphate backbone reactions in the major groove
Specific interactions in the minor groove
Specific interactions in the major groover
EcoRI looks for 2 G-C on top of each other because they're more bendable
Where in the GAATTC sequence does EcoR1 cut?
After the C, before the C
After the A & T
In the middle
Eco RI needs an Fe iron to bind
Where are the distortions of the DNA caused by EcoRI?
In the centre of GAATTC
G/AATT/C
GA/AT/TC
Hydrophobic interactions in ECORI-DNA ineractions recognise what?
AA atacking
AT stacking
TT stacking
ECORI uses water mediated interactions to assist DNA recognition
What is strange about the EcoRV restriction enzyme?
It has an abnormally large binding site
It leaves blunt ends
It binds to 2 different sequences
EcoRV forms a dimer
ECORV creates major distortions where?
At the ends
One residue in
ECORV binds to the backbone in the minor groove and wraps 'arms' around into the major groove
The recognition loops in the major groove cause...
H bonding between stacks
H bonding between A-G
H bonding between T-C
ECORV looks for ATAT bases
Where in the GCGC sequence does M.Hal methylate?
The first G
The first C
The second G
The last C
Methylation prevents the restriction enzyme from recognizing a sequence, so it cannot be cot
Describe the actions of methylases
Methylate both strands at once
Methylate one strand at a time
Are monomers
Are dimers
Interactions 'pull out' one of the bases to gain chemical access
What is the donor for methylation?
S-adenosyl-methionine
Methanol
Methide
How the methylation interaction studied?
The cystine had the 5th H replaced with an F
Trapping it in the active site
The cystine had the 3rd H replaced with an F
Once in the active site, a thiol bond forms between the cystine and the cytosine so the methyl can be added
What is inserted to bind with the widowed G?
Arg
Cys
The inserted bases fill up the space left by the flipped out cytosine
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