DNA Structure Flashcards
What did Maurice Wilkins and Rosaline Franklin find?
Experimental evidence that DNA = a helix
They were taking DNA in solution - adding ethanol at various ratios - then pulling out fibres of DNA from the liquid
Then putting these fibre into an intense X-ray beam - and following diffraction if the X-ray beam by the DNA
the crosswires of the diffractions the middle = show its a helix
They found different forms of helix
Base pairs on inside & sugars and phosphates on outside
Maurice Wilkins - using diff levels of ethanol- produce different form of DNA = A form
= right handed DNA helix - more tightly wound - size of minor & major grooves = close (not much different to B from (predominant form)
Z DNA = left handed coil DNA - found inside cells ( function not established well)
What are the 2 polynucleotide strands that make up the 2 chains of DNA in the helix composed of?
Composed of Nucleotides ( repeating base, deoxyribose sugar, phosphate) linked by 3’-5- phosphodiester bonds
What is the DNA helix held together by?
Held together by hydrogen bonds between AT & CG bonds
When you look at DNA from a distance - the amt of A = roughly amount of T and same w C & G
CG base pairs
Cytosine - Pyrimidine (single six membered ring)
Guanine - Purine - 2 runts
Form 3 hydrogen bonds between them
AT base pairs
Thymine - six membered & pyrimidine
Adenine - purine
Only 2 hydrogen bonds - not as strong of an interaction of CG base pair
When AT & CG packed together in DNA = form very smooth and uniform helix - as both similarly sized
Feature of a DNA helix
2 anti parallel polynucleotide chains for a right handed helix
Bases on inside of helix-phosphate & sugars on outside
Polynucleotide chains held together by Hydrogen bonds between AT & CG
One strand = complementary to other strand
What structure forms an A from helix?
Transfer RNA ( tRNA )
Single RNA chain - has a lot of self complementarity - so can fall back in itself to form areas or A form helix
What are some unusual DNA structures?
Left handed / Z-DNA
Formed by 5’…GCGCGCGC or 5’GTGTGTGT
alternating purine - pyrimidine sequence
Four stranded Junction = Holliday Junction
Tetraplex DNA = 4 stranded DNA helix - formed at the end of chromosomes - the telomeres of chromosomes - involves G rich DNA sequences
What are Holliday Junctions?
Arise in living cells through
DNA strand exchange between 2 homologous chromosomes - red & blue duplexes
Important role in DNA repair & exchange of genetic information
What are the levels of DNA structure?
Primary = Sequence of bases ( determined by DNA sequencing methods)
Secondary = Helical structure ( can be probed by X-ray techniques & chemistry)
Tertiary = DNA supercoiling - DNA helix urself = coiled in space ( can use electron microscopy)
Quaternary = Individual chromosomes that are interlocked.
Produced naturally in bacteria - at end of a round of replication -
where bacterial DNA = circular - the 2 daughter chromosomes are interlocked like 2 locks in a chain
What can you infer from gene sequence?
What the protein sequence must be
What is the most widely used method of DNA sequencing?
Sanger Sequencing
DNA strand = copied w DNA polymerase - in presence of inhibitors that arrest DNA synthesis - specifically at A,C,G,T
The DNA strands = separated by length on a polyacrylamide gel
If the DNA or incorporated inhibitor = radioactive or fluorescent - the DNA bands can be visualised & the sequence read
700-1000 bases per read
Why is having DNA sequence so important?
Important to understand gene function
Identify anti-microbial or anti-cancer targets or vaccine candidates
Describe bacterial / eukaryotic DNA
In most species - bacterial DNA = Circular ( e.coli= 3x106bp roughly)
Supercoiled in the cell - the DNA helix = twisted in space
Supercoiling caused by enzyme DNA Gyrase
What is DNA Gyrase?
An enzyme that uses ATP - as an energy source - and takes a relaxed DNA circle & converts it into a supercoiled space
Why is supercoiling important?
It is very important for facilitating bacterial DNA replication
An essential enzyme in bacteria
What do the enzymes Topo IV & Topo I ( related to Gyrase) do?
Reverse reaction to Gyrase
Take out the supercoiled & allows DNA to become relaxed
In the bacterial cell - there is competition between those activities - sets the level of supercoiling at some intermediate level
The size of bacteria is small compared to the size of its DNA, what happens?
The DNA has to be further compacted than what the supercoiling gives
E.g the E.coli chromosome = organised into 50 supercoiled loops so the DNA can fit inside
What is a karyotype analysis?
Human DNA = 23 pairs chromosomes. Can be visualised using this
Where you arrest cells at mitosis - then break the nucleus & spread out all the chromosomes & label them w particular pieces of DNA - which Dye is attached
So you can identify the individual chromosomes and sort then out
Far more DNA in a eukaryotic cell than in a bacterium
In the chromosomes, the DNA is complexed w histones. What are histone?
Basic proteins which interact w the negative charge in the phosphates in DNA and allow the DNA to be packaged
What is chromatin?
There are also other proteins (as well as histones) involved in binding DNA
This whole nucleoprotein complex = called Chromatin
What is the foundation of chromatin?
Nucleosome ( the building block)
First seen in the electron microscope
The DNA - seemed to be present when it came out of the nucleus as a ‘ BEADS ON A STRING’ structure
Regularly spaced nucleosomes interacting w DNA
DNA = string Nucleosomes = beads
Structure of nucleosomes
Determined by X-ray crystallography
Made up of several different proteins & DNA
In the middle = 4 different histone types = histones 2a, 2b, 3 and 4
Highly basic, small proteins - able to interact w the negative charges from the phosphates of the DNA
This core = sometimes called the Histone Aptamer = because there are 2 copies of each of the 4 histones
The DNA = coiled on the outside of the histone core
Roughly 2 super helical turned of the DNA = wrapped outside the nucleosome
Core DNA = DNA - physically associated w the histones
Linker DNA = DNA - runs between adjacent histones
