Nucleotides and nucleic acids I Biochemistry 302 January 18, 2006
Nucleotides and nucleic acids IBiochemistry 302
January 18, 2006
http://biochem.uvm.edu/courses/kelm/302
User: studentPW: nucleicacids
Central Dogma of Molecular Biology(Cell as a factory analogy)
DNA = permanent repository which stores master plans
RNA = temporary repository copyof certain plans Working RNAs (e.g.
rRNA, snRNA). Adapter RNAs (e.g.
tRNA, miRNA) Intermediary RNAs (e.g.
mRNA). Protein = working
machineryFig. 4.23
Basic chemical structure of DNA and RNA (heteropolymers of nucleotides)
Monomer composition (nucleotide) heterocyclic
pentose sugar phosphate nitrogenous base
RNA: polar ribose phosphate backbone
DNA: polar deoxyribose phosphate backbone (no 2-hydroxyl)
Nucleotides joined by 3,5- phosphodiester linkages
Nitrogenous bases side chains
Lehninger Principles of Biochemistry, 4th ed., Ch 8
Major nitrogenous bases found in DNA and/or RNA (purines & pyrimidines)
DNA: A, G, C, T RNA: A, G, C, U N--glycosyl bond: 1
carbon of ribose and N9 of Pur base (A, G) or N1 of Pyr base (C, T, U)
Pur or Pyr base + ribose = nucleoside
Fig. 4.2
parent compounds
Nucleotide Nomenclature
DNA
RNA
Chemistry of nucleotide components Phosphate group
Strong acid pKa ~1 for primary ionization,
~6 for secondary
Purine/Pyrimidine (pKa ~2.4-9.5) Weak tautomeric bases
Isomers differing in position of H atoms & double bond.
Less stable imino & enol forms found in special base interactions.
Conjugated double-bonds Resonance among ring atoms Absorb UV light
Fig. 4.4Lehninger Principles of Biochemistry, 4th ed., Ch 8
Chemical stability of polynucleotides(contribution of the ribose ring)
Hydrolysis of DNA and RNA is thermodynamically favorable but very slow.
Acid-labile bond (purine glycosidic linkage in DNA but not RNA
Base-labile bond (PDE bond in RNA but not DNA)
Nucleases (endo & exo, specific & non-specific) promote rapid hydrolysis of PDE bonds in DNA or RNA.
Dehydration-resistant (e.g. DNA in fossils) but water content (level of hydration) affects secondary structure
Lehninger Principles of Biochemistry, 4th ed., Ch 8
DNA and genetics: a historical perspective ~1868 Friedrich Miescher isolates phosphorus-containing
substance nuclein from nuclei of leukocytes and salmon sperm, noted 2 portions Acidic (DNA), Basic (Protein)
CW 1860s to 1940s Genetic inheritance dictated by proteins Nucleic acid too simple (4 nucleotides vs ~20 amino acids DNA merely a structural material present in the cell nucleus.
1944 to 1952 DNA transfer & labeling studies point to DNA as the repository of genetic information.
Late 1940s Chargaffs rules of DNA composition A = T; G = C; A + G (purines) = C + T (pyrimidines)
1953 Watson & Crick propose structure of DNA.
Hershey-Chase, 1952Avery, MacLeod, and McCarty, 1944
T2 bacteriophage infection
Viral T2 32P-DNA (not 35S-protein) transferred to and propagated in E. coli
Elucidation of DNA structureFranklin and Wilkins 1953; Kings CollegeWatson and Crick 1953; Cambridge Univ.
R. Franklin & M. Wilkins X-ray diffraction pattern of wet DNA fibers consistent with regular, repetitive helical 3D structure w/ 2 distinct periodicities. Primary repeat ( 3.4 ) Secondary repeat (34 )
J. Watson & F. Crick Built best fit model based on X-ray data, Chargaffs rules, DNA chemical composition, & clever deduction. Ten residues/turn (34 ) Helical rise (3.4 , distance betw
vertically stacked bases Two DNA strands/helix (fiber
density)R. E. Franklin and R. Gosling (1953) Nature 171:740
Cross pattern typical of helix
Properties of nucleotide bases 3D structure of nucleic acid pH-dependent tautomers
Adenine and Cytosine (amino form at pH 7) Guanine and Thymine (keto form at pH 7)
Functional groups (H-bonding) Ring nitrogens Carbonyl groups Exocyclic amino groups
Highly conjugated resonance Pyrimidines (planar) Purines (nearly planar slight pucker)
Hydrophobic character Hydrophobic stacking interactions van der Waals interactions between
uncharged atoms
Watson and Crick 1953Intuition: H-bonding between certain baseson opposite strands stabilizes the helix
Geometric Features: H-bonding between A=T, GC base pairs distance between C-1 atoms the same constant helical diameter
Bases stacked & slightly offset inside the double helix
Deoxyribose-phosphate backbone exposed to water
Pentose ring in C-2 endoconformation (sugar pucker)
antiparallel strands
bp stackingand rotation relative to long axis
H-bonding (different # in A=T vs GC bps)
~1.08 nm
36
Rise = 0.34 nm
Fig. 4.10
H-bonding pattern in W-C base pairs and numbering convention
A = T (N6,N1) = (O4,N3)
Lehninger Principles of Biochemistry, 4th ed., Ch 8
G C(O6,N1,N2) (N4,N3,O2)
antiparallel strands
(H-bond: two electronegative atoms, such as nitrogen and oxygen,
interacting with the same hydrogen)
Other features of Watson-Crick model
Right handedness (counterclockwise rotation)
Antiparallel strands Major/minor grooves
Created by offset base pairing of 2 strands
Major groove allows direct access to bases
Minor groove faces ribose backbone
Base-pairing explains Chargaffs rule A/T or G/C ~1 in organisms with dsDNA genomes.
van der Waals radius of atoms
3
5
Fig. 4.11
Other views of the Watson-Crick model for the structure of DNA
Because B-DNA is really 10.5 bp/turn.
Lehninger Principles of Biochemistry, 4th ed., Ch 8
Ribose and phosphate oxygens are in blue.
Phosphorus atoms are in yellow. Atoms comprising
bases are in gray.
Were Watson and Crick right?
Limitations of fiber diffraction studies Fiber heterogeneity Modeling intensive
(idealized version) Enhanced precision of
crystallography Atom positions specified Structure of B-DNA more
distorted than Watson-Crick model
Bending occurs wherever 4 adenosine residues appear in a row in one strand
R.E. Dickerson et al. 1983
DNA Bending
Fig. 4-16
Secondary structural variants (deduced from fiber diffraction and crystal structures)
B-form DNA fibers prepared under
high humidity Form found in cells
A-form (compact) DNA fibers prepared under
low humidity RNA-RNA and RNA-DNA
hybrids Z-form (zigzag)
elongated left-handed DNA alternating C (or 5-meC) &
G residues in alternating anti and syn glycosyl bond conformation Each structure has 36 bp.
Z-DNA: deeper narrowminor
groove
A-DNA: deeper narrow major
groove
Properties of the three forms of DNA
Pitch = (Helix rise)(base pairs/turn)
Lehninger Principles of Biochemistry, 4th ed., Ch 8
Structural variation in DNA & dsRNAnucleotide conformation
Steric constraints restrict rotation about bonds 4 (sugar pucker) and 7 (C-1-N-glycosyl bond) in different DNA structural variants (A, B, Z).
H
Lehninger, Principles of Biochemistry, 4th ed., Ch 8
Structural variation in DNA & dsRNA-furanose or sugar pucker
B-DNA A-DNA (or RNA)
Lehninger Principles of Biochemistry, 4th ed., Ch 8
What drives B-DNA into an A-DNA conformation?
Similarities Helical sense W-C base pairing
Differences Position of bases with
respect to helical axis Base tilt Groove width and depth
(shallower minor groove in A-DNA but deeper major groove)
11 bp/turn in A-DNA Rise, pitch (repeat), and
rotation per residue are smaller in A-DNA
Fig. 4-15
No H2O