Biochem Fund
Transcript of Biochem Fund
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Donald Voet • Judith G. Voet • Charlotte W. Pratt
Fundamentals of BiochemistrySecond Edition
Chapter 3:
Nucleotides, Nucleic Acids, andGenetic Information
Copyright © 2006 by John Wiley & Sons, Inc.
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Nucleic acids
DNA, RNA
(consists of nucleotides)
Bases
SugarsPhosphates
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The bases of nucleotides
planar, aromatic, heterocyclic molecules
Cytosine
UracilThymine
Adenine
Guanine
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Sugars
Primed numbers
Dexoyribonucleic acid (DNA)
Ribonucleic acid (RNA)
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Nucleosides: base + sugar
Nucleotides: base + sugar + phosphate
glycosidic bond
phosphoester bond
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The best known nucleotide
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Nucleic acid structure
Polynucleotides: mono-, di-, oligo-
Phosphodiester bondDirection: 5’-, 3’-ends
5’-AUCG-3’
pAUCG
AUCG
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DNA double helix
1. Chargaff’s rules
A=T, G=C
GC ratio
2. Bases in tautomeric forms
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Watson-Crick model of double helix
3D-structure
two polynucleotide chains forming a double helix
Antiparallel forming right handed helix
Bases occupy the core:sugar-phosphate chain in the peripheryminimizing repulsion between the charged groups
minor and major grooves
Hydrogen bonded base pairs
complementary base pairing
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Chromosome
Genome
Gene
Diploid
Haploid
Base pairs (bp)Kilobase pairs (kb)
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RNA: single stranded molecule
double stranded RNA is possible (p824)intramolecular base-pairing: 3D structure
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Structure of yeat tRNApheRNA-DNA hybrid
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Some RNAs are catalysts: ribozymes
Self cleavage at the scissile bond
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RNA is alkali unstable
DNA is acid unstable
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Donald Voet • Judith G. Voet • Charlotte W. Pratt
Fundamentals of BiochemistrySecond Edition
Chapter 23:
Nucleic Acid Structure
Copyright © 2006 by John Wiley & Sons, Inc.
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Osmotically lysed bacteriophage T2
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In ideal B DNA
Near perfect two fold symmetry
10 bp per turn
Base planes
perpendicular to the axis
3.4A van der Waals thickness
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Several distinct structuresdepending on the solvent composition and base sequences
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Flexibility of DNA
Each base pair depending on sequences
Flexible rod
More severe distortions by protein binding
However, limited conformational flexibility
7 torsion angles determiningthe conformation of a nucleotide unit
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Glycosidic bond rotation is not free
Purine has two permissible orientations: syn and anti
Pyrimidine is stable with anti-conformation
All bases are in anti in most double helix
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Ribose ring flexibility
Ribose ring is not planarOut of plane atoms: C2’ and C3’
C3’-endo, C2’-endo: on the same side of C5’
Sugar-phosphate backbone is constrained
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Supercoiled DNA
No supercoiling tightly supercoiled
EM of supercoiled DNAs
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Supercoiling and superhelicity
Superhelix topology
L = T + W
The difference between writhing and twist
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Forces stabilizing nucleic acid structures
Denaturation and renaturation
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Hyperchromatic effectUV absorption increases ~40% upon denaturation
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denaturation & renaturation incomplete renaturation
Melting curve of DNA: Tm depends on GC ratio
rapid cooling
slow cooling
cooperativeprocess
Complete renaturation
The principle of denaturation and renaturation is important for DNA manipulation
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The principle of denaturation and renaturation is important for DNA manipulation
Polymerase Chain Reaction (PCR)
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Base pairing is essential but not enough for helix stability
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Base pairing is essential but not enough for helix stability
Base stacking and hydrophobic interactions
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Ionic interactions
Charged phosphate groups
Monovalent cationsDivalent cations: specific binding to phosphate groups
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Fractionation of nucleic acids
Chromatography
ElectrophoresisUltracentrifugation
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Intercalating agents
for DNA staining
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Eukaryotic chromosome structure
Packaging of chromosomes in a cell
23 human chromosome: 3.2 billion bp x 3.4A = 1 m
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Nucleosomes: chromatin particles
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166-bp nucleosome: role of H1
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H1 bound chromatin H1 depleted chromatin
Higher levels of chromatin organization
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Higher levels of chromatin organization
30 nm diameter chromatin filaments
Model building
EM
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Histone depleted chromosome Higher magnification
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