L2: Structure of DNA & RNA Flashcards
Primary Structure of DNA
- phosphoester bond: attaches sugar to phosphate (creates backbone)
- glycosidic bond: attaches base to sugar
- conventionally read from 5’ to 3’
Secondary structure of DNA
- Watson-Crick base pairing: conventional interaction (A:T and G:C)
- antiparallel
- major and minor groove
- can be right- or left-handed
secondary structure of DNA - handness of helix
- Depends on upward direction of overlying DNA strand:
1. points to the left, then left-handed helix
2. points to the right, then right-handed helix
secondary structure of DNA - why do the grooves differ in size
- due to the geometry of the base pairs
- angle between the glycosidic link
connecting the base to the sugar
-
secondary structure of DNA - explain the angle between the glycosidic link for the 2 grooves
- link pointing down – minor groove
- link pointing up – major groove
major and minor grooves - why is this important
- DNA-protein interactions happen on the major groove bc:
1) major groove is more physically accessible
2) major groove is more chemically informative
major and minor grooves - chemically informative
- major groove presents chemical information that is:
1) greater in quantity than the minor groove
2) unambiguous, unlike minor groove
major and minor grooves - how is it chemically informative
- proteins fit into the DNA by reading different categories on the DNA
- major grooves will show a different ‘set up’ depending on the order of bases while a minor groove may continue to be the same
major and minor grooves: chemically informative - what are the different categories
- Hydrogen bond donor (+)
- Non-polar hydrogen
- Hydrogen bond acceptor (-)
- Methyl group
major and minor grooves: chemically informative - different categories for A-T within the major groove
- from left to right:
1. Hydrogen bond acceptor
2. Hydrogen bond donor
3. Hydrogen bond acceptor
4. Methyl group
major and minor grooves: chemically informative - different categories for A-T within the minor groove
- from left to right
1. Hydrogen bond acceptor
2. Non-polar hydrogen
3. Hydrogen bond acceptor
major and minor grooves: chemically informative - different categories for T-A within the major groove
- from left to right:
1. Methyl group
2. Hydrogen bond acceptor
3. Hydrogen bond donor
4. Hydrogen bond acceptor - different from A-T
major and minor grooves: chemically informative - different categories for T-A within the minor groove
- from left to right
1. Hydrogen bond acceptor
2. Non-polar hydrogen
3. Hydrogen bond acceptor - same as A-T
major and minor grooves: chemically informative - different categories for G-C within the major groove
- from left to right:
1. Hydrogen bond acceptor
2. Hydrogen bond acceptor
3. Hydrogen bond donor
4. Non-polar hydrogen
major and minor grooves: chemically informative - different categories for G-C within the minor groove
- from left to right:
1. Hydrogen bond acceptor
2. Hydrogen bond donor
3. Hydrogen bond acceptor
major and minor grooves: chemically informative - different categories for C-G within the major groove
- from left to right:
1. Non-polar hydrogen
2. Hydrogen bond donor
3. Hydrogen bond acceptor
4. Hydrogen bond acceptor - different from G-C
major and minor grooves: chemically informative - different categories for C-G within the minor groove
- from left to right:
1. Hydrogen bond acceptor
2. Hydrogen bond donor
3. Hydrogen bond acceptor - same as G-C
explain DNA strand denaturation
- DNA can undergo reversible strand separation which is needed for replication
- heating up denatures it
- cooling allows it to go back together (renaturation)
DNA strand denaturation - what is melting temperature (Tm)
temperature at which half the base pairs in a double stranded DNA molecule have denatured
DNA strand denaturation - what can affect melting temperature
- length of DNA molecule
- Percentage of G:C content
- ionic strength of solution
DNA strand denaturation: what affects Tm? - length of DNA molecule
- longer duplexes have a higher Tm
- bc those are held together more tightly by more base interactions
DNA strand denaturation: what affects Tm - Percentage of G:C content
- increases Tm
- due to 3 hydrogen bonds per base pair (as opposed to 2 in A:T)
- G:C has stronger stacking interactions with neighboring base pairs
DNA strand denaturation: what affects Tm - ionic strength of solution
- increases Tm
- negatively charged phosphates of backbone repel each other across two DNA strands (promotes lower Tm)
- cations shield and neutralize these charges and stabilize the helix (stops it from pulling against each other)
what form is DNA found in?
- linear in eukaryotes
- circular in prokaryotes and in plasmids
circular DNA - what is cccDNA
- covalently closed circular DNA
- it is supercoiled DNA
DNA topology - Linking number (Lk)
- number of times one strand would have to be passed through the other to completely separate the two strands
- will not change even if it is supercoiling
DNA topology - what is the Lk equation
- Lk = Tw + Wr
- Tw: Twist number
- Wr: writhe number
DNA topology - what is the twist number
- number of times one DNA strand wraps around other
- Tw = Lk in the absence of supercoiling (because Wr = 0)
DNA topology - what is the writhe number
- torsional stress that causes entire double helix to cross over itself
- has two types
DNA topology - what are the two types of writhing
- interwound or plectonemic: DNA coils in a helical fashion
- toroid or spiral: DNA wraps in a cylindrical fashion
DNA topology - toroid or spiral writhing
- Needs something to wrap around or it becomes unstable
- Ex: nucleosome (DNA wrapped around histones)
DNA topology - when is Lk negative or positive?
- negative when the DNA is left-handed
- positive value when the DNA is right-handed
DNA topology - when is Wr negative or positive (for interwound helices)?
- negative when DNA is right-handed
- positive when DNA is left-handed
DNA topology: interwound helix Wr negative and positive values - why is it like this?
- DNA is normally a right-handed helix
- if it forms a right-handed interwound helix, the DNA must twist to accommodate the writhe
- then Wr is given negative value (negatively supercoiled)
DNA topology - when is Wr negative or positive (for toroid/spiral helices)?
- negative when left-handed
- positive when right-handed
DNA topology: spiral helix Wr negative and positive - why is this important
nucleosomes can add negative or positive supercoiling
explain negative supercoiling
- DNA in cells exist in this state
- can be viewed as a storage of free energy
- allows strand separation to occur more easily
what is topoisomerase
- an enzyme that controls DNA topology
- it changes DNA linking number by introducing transient sting-stranded or double stranded breaks into the DNA
- will reseal afterwards
explain the structure and versatility of RNA
- different types:
1. transfer RNA
2. ribosomal RNA
3. messenger RNA
4. micro RNA
Structure and versatility of RNA - secondary strucuture
- RNA is capable of folding back on itself to form local regions of double helix
RNA secondary structure - RNA-protein interaction example
- Ricin
- highly toxic protein
- binds and alters the RNA of a large eukaryotic ribosome
what are ribozymes
- RNA enzymes
- they act as enzymes and can have active sites and binding sites for substrate and co-factors
- its active site (peptidyltransferase center) is composed of RNA and not protein
- this thus supports an early RNA world from which protein-based life arose
ribozymes - how does an enzyme work
- it binds to a substrate to facilitate a chemical reaction
- it then releases the product and repeats
