Lecture 3: DNA Structure and Recognition Flashcards
(27 cards)
What are the building blocks of DNA and RNA?
- Both have phosphoribose backbone
- deoxyribose (prime 2 has OH) or ribose (prime 2 has H)
- phosphate
Bases:
- Pyramidines - 6 membered rings
- Cytosine
- Thymine - special feature of DNA
- Uracil
- Purines - combination of 5 and 6 membered rings
- Guanine
- Adenine
How are the components structured together?
- One prime end - glycosidic bond of the sugar to the bases
- Two prime end - either OH or H
- Three prime end - anchor for polymerization
- Five prime end - makes bonds with the phosphate group
To connect to another nucleotide it will form phosphodiester bond
How do 5’ and 3’ end look like?
- 3’ = free hydroxyl group
- sugar and phosphate connecting the nucleotides = backbone (they differ on the bases)
- 5’ = free phosphate group
What is meant by hybridization? What characterize it (e.g. pairs)?
= process by which hydrogen bonds form between strands of nucleotides (bases)
- could be either within one nucleotide, but more often it is 2 separate strands
- G-C form 3 hydrogen bridges
- A-T form 2 hydrogen bridges
= Watson-Crick Base pair
- They run in anti-parallel fashion - where one strand starts with 3’ the other ends with 5’
NOTE: there are also Non-Watson-Crick base pairing - which doesn’t happen very often
What then builds up the DNA double-helix?
- The orange part = backbone of phosphates and sugar groups (negatively charged - hydrophilic)
- Base pairs laying as plates in the middle
-> held together by hydrogen bonds between the bases and pi-pi stacking of those bases (“sandwich” pattern)
Look at the major and minor grooves within the double helix structure:
Major groove - have more accessible bases -> important for transcription
What are possible DNA conformations?
1) B-DNA - normal
2) A-DNA - happen under certain conditions e.g. salt
- has inner hole - the bases are further away
- major and minor grooves closer together
3) Z-DNA
- bases out of center (bases more exposed - more fragile , backbones on the other side)
Look at specific measures about DNA:
What is the main difference between A-DNA and B-DNA?
B-DNA has major and minor grooves similar in depth while A-DNA has major goove as deeper than minor and pi stacking is deviating from the center
Picture is of B-DNA
What does this picture tell us?
- Edges of major groove bases are wider than minor groove -> due to assymetrical attachment of the bases to the phosphate group
- Also notice that each side (major/minor) and each pair (A/T x G/C) show a different pattern of possible bonding (e.g. H-acceptor, hydrophobic interactions…) => specific recognition pattern for major and minor (less surface. less flexibilty)
- the pattern more complex in major grooves
What happens when bacterophages infect a bacterium?
- Lytic cycle = release new phage
- entering bacterium -> integration of its DNA into the host -> can either continue this cycle -> producing more of its proteins to build up more bacterophages -> once accumalate they can be released
- Lysogenic cycle = integrated within host
-> OR enters this cycle -> starts copying its DNA into host -> leaves it there -> if cell divides it will still have the phages DNA within it -> after many divisions it can go into lytic cycle
How is the lytic-to-lysogenic decision made? (elaborate on the two options)
Picture A
- Left side depicts strand of repressor gene that codes for repressor protein
- Right side houses Cro gene and early lytic genes
- RNA-Polymerase is facing left due to repressor proteins blocking the OR1 and OR2 segments -> will be reading one only repressor genes
- positive feedback - more repressor proteins will block the right side more
Picture B
- RNA-polymerase faces right -> reads Cro genes -> Cro proteins block OR3 (=> lytic cycle)
How can they switch?
- E.g. UV irradiation Rec A (proteases) cleaves the repressor -> RNA-polymerase goes to right (it is more beneficial to become phage and escape to new host than being killed by radiation)
Just remember:
DNA is very specific
- often only small to no sequence variation tolerated
- frequent polindromic base pairs = sequence that reads the same from forward and backward
- The non-green can vary while the green cannot
Look at the structure of the proteins mentioned:
- Notice one is alpha heliz and beta sheet structure while the other is made up by only alpha helices
But usually occurs as dimers - and if we look at the distance between a3-helices -> they are exactly one whole turn of DNA away = recognition helix
- palindromic sequence is needed so that a3-helices can lay within the major groove
- a3-helices make contact with the DNA with glutamines G28 and Q29 (to be able to read it)
- The interaction happens with BASES - grants the specificity
Bonds with DNA are usually specific but here and there we also get non-specific interactions - what does that mean?
Specific interactions are made between a protein and base pairs - it matters where the protein binds
Non-specific are made between protein and backbone of the DNA, often with the use of NH backbone of the polypeptide
- BUT here it doesn’t matter to what phosphate/part of the DNA does it bind
Look at the summary of affinity differences in DNA related to Cro protein -> overall affinity
What is the Central Dogma? Do we haveonly one type of RNA?
DNA is replicated (DNA-Polumerases) -> transcribed into mRNA (RNA-Polymerases) -> translated into a protein (ribosomes)
- Only about 2% protein coding genes from the entire Genome pool
-> the rest (about 70%) are transcribed into non-coding RNA (ncRNA) that do NOT lead to proteins
- short ncRNA and long ncRNA
See some examples of short ncRNA.
We may know siRNA and miRNA - regulation adn degradation of mRNA
- we have more info on short rather then long ncRNA
How does transcription work?
- DNA unwinds and separates into 2 strands:
- “Coding strand” = inactive strand
- “Non-coding strand” = template strand
-> RNA polymerase will read over the template strand (going from 3’ to 5’) and will synthesize complimentary strand (copy og inactive strand)
-> after RNA has passed, DNA winds up again
So what are the major steps of transcription?
1.Initiation
2. Elongation
- includes proof reading - polymerase checks whether it inserted the correct nucleotide -> if not exonucleotide cleaves out the incorrect and inserts a new one
3. Termination
- each of these steps depends on many different protein complexes
How does translation take place?
- Happens in ribosomes -> forms the peptide bonds (most antibiotics inhibit bacterial RNA)
-RNA is read by triplets i.e. one nucleotide triplet codes for one amino acid- messenger RNA is translated (mRNA)
- transfer RNA is bringing amino acids to triplets (tRNA)
How do we read the following sun of genetic code? Are there some special codes? Or codes more susceptible to mutations?
- We start in the middle (e.g.G) -> move out of the circle (e.g. G -> G) => glycine
- some amino acids can be translated from all kinds of triplets X some are too specific e.g. tryptophan (if mutation occurs at any point -> leads to a different amino acid = less protected)
- Few codes are special
- AUG = Met (builds methianine) which starts the coding process
- UAA, UAG, or UGA code for stop code
