Copyright © 2007 by W. H. Freeman and Company Berg Tymoczko Stryer Biochemistry Sixth 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 Functions 1.Provide energy for reactions: ATP ADP + P i ATP AMP + PP i 2.High energy carrier molecules UDP-glucose and CDP-diacylglycerol 3.Substrates for making DNA and RNA All NTPs and dNTPs 4.Regulatory molecules c-AMP and c-GMP 5.Coenzyme components NAD + 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 acid Cytosine cytidine cytidine 5′- monophosphate or CMP or 5′- cytidylic acid Uracil uridine uridine 5′- monophosphate or UMP or 5′- uridylic acid Thymine 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.

B-DNA DNA can exist in A, B or Z forms. B is most common, A is at low humidity, Z is left handed Minor groove = 12 A Major groove = 22 A Diameter = 20 A

H-Bonding in dsDNA

Chargaff’s Rules Purines = Pyrimidines

Overlap of bases in dsDNA

Meselson & Stahl Experiment Beginning with DNA that contains all N 15 isotope.

Semiconservative Replication of DNA N 15 -N 15 N 15 -N 14 N 14 -N 15 N 15 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 15 N 15 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 14 N 14 -N 15

Meselson & Stahl Results Replication using NTPs that are all N 14 isotope.

Detection of DNA by absorbance of aromatic bases at 260 nm (Denaturing causes an increase in A 260 )

DNA Melting (strand dissociation) Tm = 69 o (%G + %C)

Determine %G, C, A & T from Tm Tm = 69 o (%G + %C) Tm for 100% A:T dsDNA = 69 o Tm for 100% G:C dsDNA = 110 o Assume Tm was determined to be 98 o C. 98 = (%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 dsDNA Supercoiled

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 strand Coding 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 ( bases).

Reverse Transcription

t-RNA T  C loop DHU loop

t-RNA 3′-end of t-RNA

Formyl-Met fMet bonds to the AUG codon and initiates translation.

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…..AG3 termini and a splicing start..PyUPuAPy…

Recombination Exon shuffling

End of Chapter 4 Copyright © 2007 by W. H. Freeman and Company Berg Tymoczko Stryer Biochemistry Sixth Edition