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BCH5413 - 1 EXAM PREP WITH COMPLETE SOLUTION
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BCH5413 - 1 EXAM PREP WITHCOMPLETE SOLUTION
DNA structure - =- nucleic acid bases (purine (A,G) and pyrimidine (C,U,T))
- sugars (ribose, deoxyribose)
- sugars linked by phosphate groups (sugar-phosphate backbone)
*form antiparallel strands
In mammilian DNA, which base has various forms - =cytosine
Chargaff's rule - =A=T and C=G
Is DNA conservative or semi-conservative? - =semi-conservative
- parental sequence and daughter strand bind together
Alternative Double stranded DNA conformation
- A and Z DNA - =A DNA
- dehydrated form
- right handed helix
- 11 bp per helical
- bp tilted 19 degrees
- major groove narrow and deep
- found with dsRNA and RNA-DNA duplex
z-DNA
- >1% of cellular DNA
,- favored by G-C repeats and alternating purines-pyrimidines
- left handed helix
- 12 bp per turn
- bp tilted 9 degrees
- major groove flattened, nearly gone
- bound by poxvirus E3L virulence factor and down regulates apoptosis genes
Non-duplex DNA structures
- cruciform DNA - =- inverted repeat sequence
- favored by excessive negative supercoiling
- AT-rich cruciforms associated with 'fragile' DNA
Alternative modes of base pairing - =Watson Crick:
A--T and C---G
- adenine is at anti-conformation
Hoogsteen:
- adenine or guanine can be flipped to sin-conformation (different H-bonding)
Hoogsteen base pairing
- when can it occur - =- in triplex and quadruplex DNA structures
Triplex DNA - =- pyrimidine rich strand
- negative supercoiling
Chair DNA - =- two G-rich strands
- down regulation of c-myc transcription
,Intrinsic Bends in DNA - =- distortion of the ideal B-DNA conformation resulting from base
stacking in the nucleotide sequence
Eg. Adenine-tract DNA results in 20 degree bends
- also can be found in duplex-oligonucleotide model DNA
- causes 23 degree bend and mis-stacking of one GC
- DNA is not a uniform structure
Does B-DNA have a perfect structure? - =No
- sequence specific local variation in twist between paired bases, sugat conformation, tilt of
base pairs, rise distance, etc.
Supercoiled DNA
- negative v. positive - =- left handed under-twisted DNA is in a negative supercoil
- right handed over-twisted DNA is in a positive supercoil
*most is negatively supercoiled (topoisomerase releaves supercoiling)
Denaturation/Melting of DNA - =- separation of DNA strands
- can happen due to temp, salt concentration, G---C content, and length of DNA
(Hyperchromicity effect)
- as temp increases, absorption of light increases due to bases becoming less shielded when
they melt apart
The (greater/lower) the G+C content and the (higher/lower) the salt, the more stable the DNA
duplex - =greater G+C and higher salt
, What are higher stringency conditions - =high heat and low salt
What else can melt DNA? - =pH
DNA renaturing and hybridization - =- add complementary RNA and re-anneal by gradually
lowering temp.
Where are chemical groups present in the DNA - =major and minor groove
How do sequence specific DNA binding proteins bind in a sequence specific manner - =the
amino acids of sequence specific DNA binding proteins recognize the pattern of chemical
groups in the major/minor groove
- may interact through different bonding to permit sequence specific recognition
Zinc finger binding proteins - =- custom designed DNA binding proteins
- two beta sheets attached to an alpha helix coordinate by a zinc molecule
- triple zinc finger can bind in the major groove of DNA
Zinc finger custom design - =swapping the alpha helix domain and beta-hairpin segments allows
for different sequence specificity
*alpha helix domain determines sequence specificity
Zinc finger properties - =Crystallographic studies
- a single zinc finger protein can recognize 3bp of DNA
Zinc finger and DNA binding specifics - =- AA of zinc finger at position 6 recognizes first
nucleotide of DNA
- AA of zinc finger at position 3 recognizes second nucleotide of DNA
- AA of zinc finger at position -1 recognizes third nucleotide of DNA
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