Copyright © 2007 by W. H. Freeman and Company

Berg • Tymoczko • Stryer

BiochemistrySixth Edition

Chapter 4:DNA, RNA, and the Flow of

Genetic Information

Purine & Pyrimidine Bases

Ribose & Deoxyribose

Note: numbers are primed on sugars but not bases

Adenosine, a Nucleoside (base + sugar)

Anti form

ATP, a 5′-Nucleotide (base + sugar + phosphate)

Nucleotide Functions1. Provide energy for reactions:

ATP < === > ADP + Pi

ATP < === > AMP + PPi

2. High energy carrier moleculesUDP-glucose and CDP-diacylglycerol

3. Substrates for making DNA and RNAAll NTPs and dNTPs

4. Regulatory moleculesc-AMP and c-GMP

5. Coenzyme componentsNAD+ and NADP+ ; FMN and FAD

Nucleotide Carrier

Nucleotides

NAD+ (NADP+)

A 3′-nucleotide

Bases, Nucleosides, Nucleotides

Base nucleoside 5′-nucleotide

Adenine adenosine adenosine 5′- monophosphate or AMP or 5′- adenylic acid

Guanine guanosine guanosine 5′- monophosphate or GMP or 5′- guanylic acidCytosine cytidine cytidine 5′- monophosphate or CMP or 5′- cytidylic acidUracil uridine uridine 5′- monophosphate or UMP or 5′- uridylic acidThymine thymidine thymidine 5′- monophosphate(deoxy) or dTMP or 5′- thymidylic acid

DNA & RNA Polymers

Shorthand Notation for DNA

• Deoxyribose is the vertical line. The anomeric C is #1 and phosphodiester bonds link the sugars 3’-5’.

• a and b are cleavage sites for nucleases.

P PP P OHP

A G T A C

1'

3'

5' b

a

B-DNA

DNA can exist inA, B or Z forms. B is most common, A is at low humidity, Z is left handed

Minor groove = 12 AMajor groove = 22 ADiameter = 20 A

H-Bonding in dsDNA

Chargaff’s Rules

Purines = Pyrimidines

Overlap of bases in dsDNA

Meselson & Stahl

Experiment

Beginning withDNA that containsall N15 isotope.

Semiconservative Replication

of DNA

N15-N15

N15-N14 N14-N15

N15-N14 N14-N14 N14-N14 N14-N15

N15-N14 N14-N14 N14-N14 N14-N14 N14-N14 N14-N14 N14-N14 N14-N15

Meselson & Stahl

Results

Replication usingNTPs that areall N14 isotope.

Detection of DNA by

absorbance of aromatic bases at 260 nm

(Denaturing causesan increase in A260)

DNA Melting (strand dissociation)

Tm = 69o + 0.41(%G + %C)

Determine %G, C, A & T from Tm

Tm = 69o + 0.41(%G + %C)

Tm for 100% A:T dsDNA = 69o

Tm for 100% G:C dsDNA = 110o

Assume Tm was determined to be 98oC.

98 = 69 + 0.41(%G + %C)29 = 0.41(%G + %C)29/0.41 = 70.7 = (%G + %C) %G = %C = 70.7/2 = 35.35% %A = %T = 29.3/2 = 14.65%

Circular Mitochondrial dsDNA

Relaxed

Circular Mitochondrial dsDNASupercoiled

DNA Synthesis

Addition of an AMP residue.DNA synthesis occurs from 5′ to 3′

DNA Synthesis

Addition of a GMP residue

DNA Coupling Mechanism

Common Types of RNA

r-RNA, t-RNA & m-RNA

Transcription

Synthesis of m-RNA

Template strand = antisense strandCoding strand = sense strand

Promotor region for m-RNA

Region of DNA that controls the start of m-RNA synthesis.

Stem-loop structure in RNA

Simple stem-loop structure in RNA.Shows short segments of base pairing in RNA.

Note the poly-U structure at the 3′ end of Ecoli m-RNA.

Eucaryotic m-RNA

The 5′ end is capped with 7-methylG through a -ppp- bridge. Other caps are known.The 3′ end has a poly-A tail (100-200 bases).

Reverse Transcription

t-RNA

TC loopDHU loop

t-RNA3′-end of t-RNA

Formyl-Met

fMet bonds to the AUG codon and initiates translation.

Translation Start

Translation Start

Splicing m-RNA

Introns = intervening sequences (excised), vary in size & number

Exons = expressed sequences

Eucaryotes have split genes

m-RNA with no introns

Hybridization to genomic DNA is smooth

m-RNA with introns

Hybridization to genomic DNA is looped

Intron consensus sequence

All introns have conserved 5GU…..AG3termini and a splicing start ..PyUPuAPy…

RecombinationExon shuffling

End of Chapter 4

Copyright © 2007 by W. H. Freeman and Company

Berg • Tymoczko • Stryer

BiochemistrySixth Edition