• Chapter 17

• Transcription, RNA
Processing, and
Translation

Copyright © 2019 Pearson Canada Inc. 17 - 1

Chapter 17 Opening Roadmap

Copyright © 2019 Pearson Canada Inc. 17 - 2

Introduction to Transcription, RNA Processing, and
Translation
• Proteins are the stuff of life

• A cell builds the proteins it needs from instructions encoded in its genome
– Transcribe DNA into RNA
– Translate mRNA into protein

Copyright © 2019 Pearson Canada Inc. 17 - 3

An Overview of Transcription
• RNA polymerases synthesize an mRNA version of the instructions stored in DNA
– Uses ribonucleoside triphosphates (NTPs)
– Matches complementary bases to one strand of DNA
– RNA is synthesized in the 5′ → 3′ direction

• Only one strand of DNA is the template

– Called the template strand
– Other strand is the non-template, or coding strand
 Matches the sequence of the mRNA (except U for T)
• Like DNA polymerases
– RNA polymerases perform template-directed synthesis in the 5′-to-3′ direction

• Unlike DNA polymerases

– RNA polymerases do not require a primer to begin transcription

• Bacteria have one RNA polymerase

• Eukaryotes have at least three distinct types:

– RNA polymerase I, II, and III

Copyright © 2019 Pearson Canada Inc. 17 - 4

Transcription Is the Synthesis of RNA from a DNA
Template

Copyright © 2019 Pearson Canada Inc. 17 - 5

Sigma Is the Promoter-Recognizing Subunit of the
Bacterial RNA Polymerase Holoenzyme

• Bacterial promoters
– Are 40−50 base pairs long
– Have a –10 box: a TATAAT sequence about 10 bases upstream of the
transcription start site
 Downstream means in the same direction that RNA polymerase moves
 Upstream is the opposite direction
– Also have a –35 box: TTGACA sequence about 35 bases upstream of the
transcription start site
Copyright © 2019 Pearson Canada Inc. 17 - 6
Events Inside the Holoenzyme
• Transcription begins when the sigma part of the holoenzyme complex binds to the –35
and –10 boxes

• Sigma can bind in only one orientation, so the orientation of the promoter determines
– Which DNA strand will be used as the template
– In which direction RNA polymerase will move
• RNA polymerase opens the DNA double
helix
– Creates a transcription bubble
– The template strand is threaded
through the RNA polymerase active
site

• Incoming NTPs enter a channel in the

enzyme and diffuse to the active site

• Complementary NTPs pair with

complementary DNA bases, and
polymerization begins

Copyright © 2019 Pearson Canada Inc. 17 - 7

Sigma Orients the DNA Template inside RNA Polymerase
• Initiation is the first phase of transcription

• RNA polymerase cannot initiate transcription on

its own
– In bacteria, sigma protein must bind to it
first
– RNA polymerase and sigma form a
holoenzyme
 RNA polymerase is the core enzyme
because it has the active site

• Sigma binds to DNA segments called

promoters that promote the start of
transcription
– Different sigma proteins bind to
promoters with slightly different DNA
base sequences
– Allows organism to activate certain
genes in response to environmental
change

Copyright © 2019 Pearson Canada Inc. 17 - 8

Transcription in Bacteria Terminates When an RNA
Hairpin Forms
• During the elongation phase of
transcription
– RNA polymerase moves along the
DNA template
– Synthesizes RNA in the 5′ → 3′
direction

• Transcription ends with termination

– RNA polymerase transcribes a
transcription-termination signal
– Codes for RNA that forms a hairpin
structure
– Causes the RNA polymerase to
separate from the RNA transcript

Copyright © 2019 Pearson Canada Inc. 17 - 9

Transcription in Eukaryotes
• Eukaryotic transcription has several differences:
– Three RNA polymerases
– More diverse promoters, including TATA box
– Basal transcription factors instead of sigma proteins
– At termination, a poly(A) signal is transcribed rather than a hairpin, and
the RNA downstream is cut
– Transcription occurs in the nucleus, and translation occurs in the
cytoplasm

mRNA Processing in Eukaryotes

• In bacteria, mRNAs are translated immediately, sometimes even before
transcription is complete

• In eukaryotes, the initial product of transcription is an immature primary

transcript or pre-mRNA

• Primary transcripts must undergo RNA processing before they can be

translated

Copyright © 2019 Pearson Canada Inc. 17 - 10

Table 17.1 Transcription, RNA Processing, and
Translation in Bacteria and Eukaryotes
Process Bacteria Eukaryotes

Transcription Blank Blank

RNA polymerase(s) One Three; each produces a different class of RNA

Promoter structure Typically contains a −35 box and a More variable; often includes a TATA box about
−10 box −30 from the transcription start site
Proteins that Sigma; different versions of sigma Many basal transcription factors
associate with bind to different promoters
promoter
RNA processing Blank Blank

mRNAs Rare Extensive; several processing steps occur in the

nucleus before the RNA is exported to the
cytoplasm: (1) enzyme-catalyzed addition of 5′
cap, (2) splicing (intron removal) by spliceosome,
(3) enzyme-catalyzed addition of 3′ poly(A) tail
tRNAs and rRNAs Cut into functional pieces if required; Cut into functional pieces if required; specific
specific bases are modified bases are modified
Translation Initiation occurs at the start codon Initiation begins at the 5′ cap and reaches the
directly; elongation and termination start codon by scanning; elongation and
similar to eukaryotes termination similar to bacteria

Copyright © 2019 Pearson Canada Inc. 17 - 11

 RNA Splicing
• Primary RNA transcripts contain exons and introns

• Introns are removed by splicing

– Catalyzed by small nuclear ribonucleoproteins, or snRNPs
– Form a complex called a spliceosome
 Most complex macromolecular machine known
 Composed of over 300 proteins

• Splicing allows different mRNAs and proteins to be produced from a single

gene

Copyright © 2019 Pearson Canada Inc. 17 - 12

Introns Are Spliced Out of the Primary Transcript

Four steps to splicing:

1. snRNPs bind to a GU at the 5′
exon–intron boundary and to an A
near the end of the intron
2. Other snRNPs form a spliceosome
3. The intron forms a lariat of a single-
stranded stem plus a loop with A as
its connecting point
4. The lariat is cut out and the two
exons are linked. The intron is
degraded

Copyright © 2019 Pearson Canada Inc. 17 - 13

 In Eukaryotes, a Cap and a Tail Are Added to mRNAs
• Primary RNA transcripts are also processed by the addition of
– A 5′ cap—a modified guanine nucleotide that enables ribosomes to bind
and protects from degradation
– A 3′ poly(A) tail—100–250 adenine nucleotides; is needed for translation
and protects from degradation

• After splicing and addition of the cap and tail, the product is a mature mRNA

• Mature mRNAs contain untranslated regions (UTRs) at both ends

Copyright © 2019 Pearson Canada Inc. 17 - 14

Ribosomes Are the Site of Protein Synthesis
• In translation, the sequence of bases in an mRNA is converted to an amino
acid sequence in a protein

• Ribosomes catalyze translation of the mRNA sequence into protein

• In bacteria, ribosomes begin translating an mRNA before transcription is

complete
– Multiple ribosomes attached to an mRNA form a polyribosome
– Many copies of a protein are produced from one mRNA

Transcription and Translation

Are Coupled and Occur
Simultaneously in Bacteria

Copyright © 2019 Pearson Canada Inc. 17 - 15

Translation in Bacteria and Eukaryotes
• In eukaryotes, transcription and translation are separated

• mRNAs are synthesized and processed in the nucleus

– Mature mRNAs are transported to the cytoplasm for translation by
ribosomes
– Polyribosomes form

Copyright © 2019 Pearson Canada Inc. 17 - 16

How Does an mRNA Codon Specify an Amino Acid?
• There were two hypotheses regarding how the amino acid sequence is coded
by a sequence of nucleotides
1. mRNA codons and amino acids interact directly
2. Crick proposed that an adapter molecule holds amino acids in place while
interacting with a codon

• The adapter molecule was later found to be a small RNA called transfer RNA
(tRNA)
– tRNAs bind to amino acids
– Transfer them to the growing polypeptide

Copyright © 2019 Pearson Canada Inc. 17 - 17

Figure 17.10 Amino Acids Are Transferred from
tRNAs to Proteins

Copyright © 2019 Pearson Canada Inc. 17 - 18

The Structure of an Aminoacyl Transfer RNA

• tRNAs are relatively short: 75–85 nucleotides long

• They can form secondary structure by folding into a stem-and-loop

• A CCA sequence at the 3′ end is the binding site for amino acids
– An aminoacyl tRNA is a tRNA linked to its amino acid

• The loop at the opposite end forms the anticodon

– Has a sequence of three nucleotides
– Can base-pair with the mRNA codon
Copyright © 2019 Pearson Canada Inc. 17 - 19
How Are Amino Acids Attached to tRNAs?
• ATP is required to attach tRNA to an amino acid

• Enzymes called aminoacyl-tRNA synthetases “charge” the tRNA by

– Catalyzing the addition of amino acids to tRNAs

• For each of the 20 amino acids

– There is a different aminoacyl tRNA synthetase
– There are one or more tRNAs

Aminoacyl-tRNA Synthetases Couple the

Appropriate Amino Acid to the Appropriate tRNA

Copyright © 2019 Pearson Canada Inc. 17 - 20

 How Many tRNAs Are There?
• There are 61 different codons but only about 40 tRNAs in most cells

• Crick proposed the wobble hypothesis

– The anticodon of tRNAs can still bind successfully
– To a codon whose third position
– Requires a nonstandard base pairing

• One tRNA is able to base-pair with more than one type of codon

Copyright © 2019 Pearson Canada Inc. 17 - 21

 The Structure of Ribosomes and Their Function in
Translation
• Ribosomes contain
– Many proteins
– Ribosomal RNA (rRNA)

• Ribosomes can be separated into

two subunits
1. The small subunit holds the
mRNA in place
2. The large subunit is where
peptide bonds form

• During translation
– Three tRNAs line up within the
ribosome

Copyright © 2019 Pearson Canada Inc. 17 - 22

The Structure of Ribosomes and Their Function in
Translation

• Three tRNAs are bound to the

corresponding codon in the mRNA

• The tRNAs fit into three sites in the

ribosome:
1. The A site is the acceptor site for
an aminoacyl tRNA
2. The P site is the peptidyl site
where a peptide bond forms
3. The E site is where tRNAs
without amino acids exit the
ribosome

Copyright © 2019 Pearson Canada Inc. 17 - 23

The Structure of Ribosomes and Their Function in
Translation
• The ribosome is a molecular machine that
synthesizes proteins in a three-step
sequence:
1. An aminoacyl tRNA carrying the
correct anticodon for the mRNA
codon enters the A site
2. A peptide bond forms between the
amino acid on the A-site tRNA and the
polypeptide on the P-site tRNA
3. The ribosome moves down the mRNA
by one codon and all three tRNAs
move down one position
 The tRNA in the E site exits
 The A site is available for another
tRNA to bind

Copyright © 2019 Pearson Canada Inc. 17 - 24

The Structure of Ribosomes and Their
Function in Translation
• The protein grows by one amino acid with each
repeat of the three steps
• Amino acids are always added to the carboxyl end
(C-terminus) of the polypeptide
• Translation has three phases:
1. Initiation
2. Elongation
3. Termination

Copyright © 2019 Pearson Canada Inc. 17 - 25

 Initiating Translation
• The initiation phase of translation begins near the AUG start codon

• The small ribosomal subunit binds to the mRNA

– At the ribosome binding site (Shine–Dalgarno sequence)
– About 6 bases upstream from the start codon
– Mediated by initiation factors

• The first tRNA is called the initiator tRNA

– It carries a modified methionine (f-Met) in bacteria

Copyright © 2019 Pearson Canada Inc. 17 - 26

Initiation Requires Binding of Initiator tRNA to
mRNA and Assembly of the Ribosome

• Translation initiation is a three-step process in bacteria:

1. The mRNA binds to a small ribosomal subunit
2. The initiator tRNA bearing f-Met binds to the start codon
3. The large ribosomal subunit binds so that the initiator tRNA is in the P site

• Translation is now ready to begin

Copyright © 2019 Pearson Canada Inc. 17 - 27
Is the Ribosome an Enzyme or a Ribozyme?
• The active site of the ribosome is entirely ribosomal RNA

• Ribosomal RNA catalyzes peptide bond formation

• The ribosome is a ribozyme

– An RNA with catalytic activity

• This supports the RNA world hypothesis

Copyright © 2019 Pearson Canada Inc. 17 - 28

 Elongation Extends the Polypeptide Chain

• Translocation occurs when the ribosome slides one

• At the start of elongation codon toward the 3′ end of the mRNA
– The initiator tRNA is in the P site – Elongation factors help move the ribosome
– The E and A sites are empty
• Translocation accomplishes three things:
• An aminoacyl tRNA binds to the codon in 1. The uncharged tRNA from the P site moves into
the A site the E site and is ejected from the ribosome
• The amino acid on the P-site tRNA is 2. The tRNA attached to the growing protein moves
transferred to the amino acid on the A-site into the P site
tRNA 3. Opens the A site to expose a new codon, which
is available to accept a new aminoacyl tRNA
Copyright © 2019 Pearson Canada Inc. 17 - 29
Elongation Extends the Polypeptide Chain

• The three steps of translocation repeat at each codon along the mRNA
1. Arrival of the aminoacyl tRNA in the A site
2. Peptide bond formation
3. Translocation

Copyright © 2019 Pearson Canada Inc. 17 - 30

Termination Occurs When the Ribosome Reaches a
Stop Codon

• Termination occurs when the A site encounters a stop codon

• A protein called a release factor enters the A site

– Resembles tRNAs in size and shape
– But does not carry an amino acid
– Hydrolyzes the bond linking the P-site tRNA to the polypeptide chain

• The newly synthesized polypeptide, tRNAs, and ribosomal subunits separate from the
mRNA
Copyright © 2019 Pearson Canada Inc. 17 - 31
The Major Steps of Gene Expression in a Eukaryotic Cell

Post-Translational Modifications
• Most proteins go through an extensive series
of processing steps
– Called post-translational modification
– Before they are completely functional

• Folding determines a protein’s shape and

function
– Molecular chaperones speed protein
folding

• Chemical Modifications
– Enzymes may add sugar or lipid groups or
phosphate groups
– addition of a phosphate group by
enzymes called protein kinases.

Copyright © 2019 Pearson Canada Inc. 17 - 32