Lecture 5 Flashcards
Primary structure:
sequence of nucleotides in the linear molecules
Secondary structure:
3D arrangement, double helix
Tertiary structure:
longer range interactions and supercoiling
X-ray diffraction of DNA:
DNA fibers are aligned along their axis. They are exposed to an X-ray beam and photographic film is positioned behind the DNA to capture diffraction of light
Diffraction theory:
it is known that a helix gives a cross-shaped pattern. DNA has 10 residues/ turn
Diffraction:
the scattering by repeating elements in the structure shows reinforcement of the scattered waves in certain specific directions and weakening in others
Constructive diffraction:
reinforce one another
Destructive diffraction:
interfere one another
Properties of diffraction wavelength:
must be shorter than the regular spacing between the elements of the structure
Short spacings in the periodic structure:
large spacings in the diffraction pattern (vice versa)
Intensities:
matter
Repeat (c):
distance parallel to the helix axis in which the structure exactly repeats itself
Reisdue (m):
some number of polymer residues
Pitch (p):
the distance parallel to the helix axis in which the helix makes 1 turn
When there is an integral number of residues:
the pitch and repeats are equal
First diffraction:
parallel to the axis of the stretched fiber
Rise (h):
the distance parallel to the axis from the level of one residue to the next (c/m)
Layer lines:
lines perpendicular to the fiber axis (inversely proportional to the repeats)
Properties of the secondary structure of DNA:
- repeat of 10 nucleotides
- pitch of 3.4 nm
- rise of 0.34 nm between two nucleotides
- stabilize by H-bonds between purines and pyrimidines
- bases are stacked by van der Waals forces
- hydrophilic phosphate backbone
- hydrophobic base pairs
Length of base pairs:
each A-T and G-C base pairing has 1.08 nm caused by purine and pyrimidine structure, each rotated at 36 degrees and 0.34 between
Chargaff’s rules:
showed that base composition varies from organism to organism: %A=%T and %G=%C
Base composition of E. coli:
40 AT, 10 GC
Base composition of human:
30 AT, 20 GC
Base composition of mycobacterium:
15 AT, 35 GC
Base composition of bacterophage:
24 A, 31.2 T, 21.5 C, 23.3 G
Most common DNA form:
B-form DNA
Properties of B-form DNA:
- backbone outside
- bases inside
- major groove
- minor groove
- antiparallel
- right-handedness
Major groove:
bases can be approached by DNA binding proteins
Atoms in DNA:
- oxygen
- carbon
- phosphorus
- carbon-oxygen-nitrogen
DNA is antiparallel:
one 5’ -OH group at the top, one 3’ P group at the bottom (vice versa),
DNA chains are held by:
hydrogen bonds between pairs of bases on opposite strands
The two DNA strands are:
complementary to one another, and a template for the replication of DNA
Conservative model of DNA replication:
one of the daughter duplexes is the conserved original duplex and one is completely new
Semi-conservative model of DNA replication:
involves unwinding the two strands and each serves as a template to copy a new strand. The daughter strands contain one of the original template strands and one new material
Dispersive model of DNA replication:
parental material is scattered through the structures of both daughter duplexes
Meselsohn and Stahl experimental data for semi-conservative replication:
- E. coli grown in 15N or 14N
- Daughter strands have a density in the middle consistent with semi-conservative replication
- Another round in light isotope shows daughter strands are solely light, consistent with semi-conservative replication
What causes A-DNA form?
- RNA-RNA double helix molecules
- DNA-RNA hybrids from helices
- DNA in low-humidity environments
The difference between B-DNA and A-DNA:
A-DNA has more bp/turn and a shorter rise between successive nucleotides
Human mitochondrial tertiary structure:
- relaxed and supercoiled allows for compaction in a cell
- circular and double stranded
E. coli motichondrial tertiary structre:
circular and double-stranded
What causes supercoiling:
wrapping around histones
Negative and positive supercoils:
overwinding will create a positive supercoil and underwinding creates a negative supercoil
Secondary structure of RNA:
- random single stranded RNA
- stacked-base structure
- “hairpin” structures
Secondary structure of viruses:
double-stranded RNA genome
Random single stranded RNA:
random coil structure of denatured single strands. There is flexibility of rotation of residues and no specific structure
Stacked-base structure:
adapted by non-self complementary single strands under “ native” conditions. Bases stack to pull the chain into a helix, but there is no H-bonding
“Hairpin” structures:
formed by self-complementary sequences (green and orange regions of the single strand); the chain folds back on itself to make a stem-loop structure
Single-stranded nucleic acid base-pairing:
forms stem-loop structures
Secondary structure of tRNA:
have extensive regions of double-stranded structures that create an overal 3D structure, important to its function
DNA denaturation:
when heated, double-stranded molecules melt into single-strands (disruption of H-bonds and van der Waals). Once cooled, they find their complementary sequences and reassociate (hybridize or anneal)
Relative strength of G-C pairs:
stronger than A/T pairs as they have 3 H-bonds instead of 2
Energy level between native DNA and denatured DNA:
denatured DNA has higher energy than the double helix because their base pairs are exposed and absorb more energy
Free energy of DNA at high temperature:
more favorable at higher temperatures and drives denaturation
Free energy of DNA at low temperature:
less favorable at lower temperatures and denaturation does not occur
Factors affecting melting temperature:
- length
- bonds
- composition
- ion solution
