2
3
1
1 = initiator proteins
2 = single strand binding proteins
3 = helicase
4 = topoisomerase (gyrase)
4
Replication
• Two DNA polymerase enzymes are
necessary for replication in E. coli
– DNA polymerase I
– DNA polymerase III
- Along each template DNA strand, leading and lagging strands can
be observed.
- The names were suggested based on synthesis at any given region.
- At any particular point in the DNA strand, if there is a leading
strand, the complementary strand will have lagging strand.
Replication
• Two DNA polymerase enzymes are
necessary for replication in E. coli
– DNA polymerase I
– DNA polymerase III
• Both have polymerase and exonuclease
activities (functions)
• First let us take a look at the polymerase
activity aspect of DNA polymerases and
then discuss exonuclease activities
Replication
• DNA Polymerase III
– Synthesize new DNA in the 5’  3’ direction
• Synthesizes long sequences of new DNA
• Is highly processive; synthesizes DNA for a long
period of time without releasing the template
• For example, synthesizes leading strand
• DNA Polymerase I
– Synthesize new DNA in the 5’  3’ direction
• Only synthesizes short sequences of new DNA
• But before it could do this, it needs to remove RNA
primers
• This is achieved by its 5’  3’ exonuclease activity
5’ 3’
exonuclease
activity of
DNA
polymerase I
Replication
• The phosphodiester backbone of
adjacent DNA fragments must be
joined after DNA synthesis by DNA
polymerases I and III
• This is done by the enzyme DNA
ligase
Both DNA polymerases have proof reading
activity
This is a 3’  5’ exonuclease activity
DNA
Polymerase
activity
Replication
• DNA Polymerase I
– Synthesize new DNA in the 5’  3’ direction
• Only synthesizes short sequences of new DNA
– 3’  5’ exonuclease activity (proofreading)
– 5’  3’ exonuclease activity (remove primers)
• DNA Polymerase III
– Synthesize new DNA in the 5’  3’ direction
• Synthesizes long sequences of new DNA
– 3’  5’ exonuclease activity (proofreading)
NOTE: DNA polymerase III does not have the
5’  3’ exonuclease activity
This week we will
complete…
Chapter 13 (transcription)
Pages 348 – 361
Chapter 15 (translation)
Pages 409 - 421
The Central Dogma
(Francis Crick, 1958)
(Transcription)
DNA

(Gene/Genotype)
(Translation)
RNA

Protein
(Phenotype)
An informational process between the genetic
material (genotype) and the protein (phenotype)
Properties of RNA
RNA has the sugar ribose rather than
deoxyribose
Properties of RNA
Nucleotides carry the bases adenine,
guanine and cytosine (like DNA)
But uracil is
found in place
of thymine
Structure of RNA
• Designate the Nucleotides
– Purines
• Guanine = G
• Adenine = A
– Pyrimidines
• Uracil = U
• Cytosine = C
Structure of RNA
Nucleotides join together,
forming a polynucleotide
chain, by phosphodiester
bonds
A phosphodiester bond
A phosphodiester bond
Usually single-stranded
Can have a much
greater variety of
complex three
dimensional
shapes than
double-stranded
DNA
Classes of RNA for
Transcription and Translation
• Informational RNA (intermediate in the process of
decoding genes into polypeptides)
– Messenger RNA (mRNA)
• Functional RNAs (never translated into proteins,
serve other roles)
– Transfer RNAs (tRNA)
• Transport amino acids to mRNA and new protein
– Ribosomal RNAs (rRNA)
• Combine with an array of proteins to form ribosomes;
platform for protein synthesis
– Small nuclear RNAs (snRNA)
• Take part in the splicing of primary transcripts in
eukaryotes
– Small cytoplasmic RNAs (scRNA)
• Direct protein traffic in eukaryotic cells
– Micro RNAs (miRNA)
• Inhibits translation and induces degradation of
complementary mRNA
RNA nucleotide sequences are
complementary to DNA molecules
New RNA is synthesized 5’ to 3’ and antiparallel to
the template
DNA template
DNA template
Adenine
Guanine
Cytosine
Thymine
Synthesized 5’ to 3’ and
antiparallel to the
template
Complementary RNA
Uracil
Cytosine
Guanine
Adenine
Only one strand of the DNA acts as a
template for transcription
The template strand can be different for different genes
But….
For each gene only one strand of DNA serve as a template
Transcription
Catalyzed by the enzyme
RNA polymerase
Single RNA polymerase (Prokaryotes)
Core enzyme
2 ,1  and 1 ’ subunits
Holoenzyme
2 , 1 , 1 ’ subunits plus
σ subunit
Polymerizes RNA
Finds initiation sites
Initiation: The region that signals the initiation of
transcription is a promoter
- 35 bases from initiation of transcription
Recognized by RNA polymerase
- 10 bases from initiation of transcription
Unwinding of DNA double helix begins here
Elongation: RNA is polymerized in 5’  3’ direction
Elongation
NTPS (ATP, GTP, CTP, UTP) are added
The energy is derived
by splitting the highenergy triphosphate
bond
Termination
RNA polymerase recognizes signals (sequence)
for chain termination
Releases the RNA and enzyme from the template
Animation on Transcription
http://highered.mcgrawhill.com/sites/0072556781/student_view0/chapter12/animation_
quiz_1.html