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replication
Protein Synthesis
The information content of DNA is in the form
of specific sequences of nucleotides along
the DNA strands
The DNA inherited by an organism leads to
specific traits by dictating the synthesis of
proteins
The process by which DNA directs protein
synthesis, gene expression includes two
stages, called transcription and translation
Transcription and Translation
Cells are governed by a cellular chain of
command
DNA RNA protein
Transcription
Is the synthesis of RNA under the direction of DNA
Produces messenger RNA (mRNA)
Translation
Is the actual synthesis of a polypeptide, which
occurs under the direction of mRNA
Occurs on ribosomes
Transcription and Translation
In prokaryotes transcription and
translation occur together
DNA
TRANSCRIPTION
mRNA
Ribosome
TRANSLATION
Polypeptide
Nuclear
envelope
TRANSCRIPTION DNA
Pre-mRNA
RNA PROCESSING
mRNA
Ribosome
TRANSLATION
(b) Eukaryotic cell. The nucleus provides a separate
Polypeptide compartment for transcription. The original RNA
transcript, called pre-mRNA, is processed in various
ways before leaving the nucleus as mRNA.
Transcription
Transcription is the DNA-
directed synthesis of RNA
RNA synthesis
Is catalyzed by RNA
polymerase, which pries the
DNA strands apart and
hooks together the RNA
nucleotides
Follows the same base-
pairing rules as DNA,
except that in RNA, uracil
substitutes for thymine
RNA
RNA is single stranded, not double stranded like DNA
RNA is short, only 1 gene long, where DNA is very long and
contains many genes
RNA uses the sugar ribose instead of deoxyribose in DNA
RNA uses the base uracil (U) instead of thymine (T) in DNA.
Table 17.1
Synthesis of an RNA Transcript
The stages of 5
Promoter
Transcription unit
3
transcription 3
Start point
DNA
5
Initiation
the polymerase initiates RNA synthesis at the
start point on the template strand.
Elongation 5
3
Template strand of
3
5
Unwound RNA
Termination DNA
DNA
transcript
2 Elongation. The polymerase moves downstream, unwinding the
DNA and elongating the RNA transcript 5 3 . In the wake of
Rewound transcription, the DNA strands re-form a double helix.
RNA
5 3
3 3 5
5
RNA
transcript
3 Termination. Eventually, the RNA
transcript is released, and the
polymerase detaches from the DNA.
5 3
3 5
5 3
Completed RNA
transcript
Synthesis of an RNA Transcript - Initiation
Promoters signal the TRANSCRIPTION DNA 1 Eukaryotic promoters
RNA PROCESSING Pre-mRNA
synthesis Polypeptide
Promoter
5 T A T A A AA 3
Transcription factors help 3 AT AT T T T 5
TATA box Start point Template
DNA strand
eukaryotic RNA 2 Several transcription
factors
polymerase recognize Transcription
factors
promoter sequences 5 3
3 5
3 Additional transcription
A crucial promoter DNA factors
sequence is called a
TATA box. RNA polymerase II
Transcription factors
5 3
3 5 5
RNA transcript
Transcription initiation complex
Synthesis of an RNA Transcript - Elongation
RNA polymerase synthesizes a single strand of RNA against the DNA
template strand (anti-sense strand), adding nucleotides to the 3 end of
the RNA chain
As RNA polymerase moves along the DNA it continues to untwist the
double helix, exposing about 10 to 20 DNA bases at a time for pairing
with RNA nucleotides
Elongation Non-template
strand of DNA
RNA nucleotides
RNA
polymerase
T C C A A
A
3
3 end
U
A E G C A
5
T A G G T T
Direction of transcription
5 Template
(downstream)
strand of DNA
Newly made
RNA
Synthesis of an RNA Transcript - Termination
Specific sequences in the DNA signal
termination of transcription
When one of these is encountered by the
polymerase, the RNA transcript is
released from the DNA and the double
helix can zip up again.
Transcription Overview
Post Termination RNA Processing
Most eukaryotic mRNAs arent ready to be translated into protein directly after being
transcribed from DNA. mRNA requires processing.
Transcription of RNA processing occur in the nucleus. After this, the messenger
RNA moves to the cytoplasm for translation.
The cell adds a protective cap to one end, and a tail of As to the other end. These
both function to protect the RNA from enzymes that would degrade
Non-coding regions may have specific chromosomal functions or have regulatory purposes
Thus, an RNA copy of a gene is converted into messenger RNA by doing 2 things:
It contains transcripts of
both introns and exons.
mRNA
Coding Introns cut out and
segment exons spliced together
Ribosome
TRANSLATION
Polypeptide
mRNA 5 Cap Poly-A tail
1 146
3 UTR 3 UTR
Figure 17.10
RNA Processing
RNA Splicing can also be carried out by spliceosomes
RNA transcript (pre-mRNA)
5
Exon 1 Intron Exon 2
Protein
1 Other proteins
snRNA
snRNPs
Spliceosome
2 5
Spliceosome
components
Cut-out
intron
3
mRNA
5
Exon 1 Exon 2
Alternative Splicing (of Exons)
How is it possible that there are millions of
human antibodies when there are only about
30,000 genes?
Alternative splicing refers to the different
ways the exons of a gene may be combined,
producing different forms of proteins within
the same gene-coding region
Alternative pre-mRNA splicing is an important
mechanism for regulating gene expression in
higher eukaryotes
RNA Processing
Proteins often have a modular architecture
consisting of discrete structural and functional
regions called domains
In many cases different exons code for the
different domains in a protein
Gene
DNA
Exon 1 Intron Exon 2 Intron Exon 3
Transcription
RNA processing
Translation
Domain 3
Domain 2
Domain 1
polypeptide Polypeptide
tRNA
Anticodon
A A A
U G G U U U G G C
5 Codons 3
mRNA
The Genetic Code
Genetic information is encoded as a sequence of nonoverlapping
base triplets, or codons
The gene determines the sequence of bases along the length of an
mRNA molecule
DNA Gene 2
molecule
Gene 1
Gene 3
DNA strand 3 5
(template) A C C A A A C C G A G T
TRANSCRIPTION
U G G U U U G G C U C A
mRNA 5 3
Codon
TRANSLATION
3
Amino acid A
C
attachment site C
A 5
C G
The anticodon is the 3 RNA bases that G C
C G
matches the 3 bases of the codon on the U G
U A
mRNA molecule A U
U C A U
* C A C AG UA A G *
G * C U C
G U G U G
*
C * C G A G
* * U C * A G G
* G AG C
(a) Two-dimensional structure. The four base-paired regions and three G C Hydrogen
loops are characteristic of all tRNAs, as is the base sequence of the U A bonds
amino acid attachment site at the 3 end. The anticodon triplet is * G
A
unique to each tRNA type. (The asterisks mark bases that have been A* C
chemically modified, a characteristic of tRNA.) * U
A G
A
Anticodon
Transfer RNA
3 dimensional tRNA molecule is roughly L shaped
5 Amino acid
attachment site
3
Hydrogen
bonds
A AG
3 5
Anticodon
Anticodon
(c) Symbol used
(b) Three-dimensional structure in the book
Ribosomes
Ribosomes facilitate the specific coupling of tRNA anticodons
with mRNA codons during protein synthesis
The 2 ribosomal subunits are constructed of proteins and RNA
molecules named ribosomal RNA or rRNA
TRANSCRIPTION DNA
mRNA
Ribosome
TRANSLATION
Polypeptide
Exit tunnel
Growing
polypeptide
tRNA
molecules
Large
subunit
E
P A
Small
subunit
5
mRNA 3
(a) Computer model of functioning ribosome. This is a model of a bacterial
ribosome, showing its overall shape. The eukaryotic ribosome is roughly
similar. A ribosomal subunit is an aggregate of ribosomal RNA molecules
and proteins.
Ribosome
The ribosome has three binding sites for tRNA
The P site
The A site
The E site P site (Peptidyl-tRNA
binding site)
A site (Aminoacyl-
tRNA binding site)
E site
(Exit site)
Large
subunit
E P A
mRNA
binding site Small
subunit
tRNA
mRNA 3
Codons
5
(c) Schematic model with mRNA and tRNA. A tRNA fits into a binding site when its anticodon base-
pairs with an mRNA codon. The P site holds the tRNA attached to the growing polypeptide. The A
site holds the tRNA carrying the next amino acid to be added to the polypeptide chain. Discharged
tRNA leaves via the E site.
Building a Molecule of tRNA
A specific enzyme called an aminoacyl-tRNA
synthetase joins each amino acid to the correct tRNA
P P P Adenosine
1 Active site binds the
amino acid and ATP.
ATP
Pyrophosphate P Pi
Pi
Pi
Phosphates
tRNA
3 Appropriate
tRNA covalently
Bonds to amino
Acid, displacing P Adenosine
AMP. AMP
4 Activated amino acid
is released by the enzyme.
Aminoacyl tRNA
(an activated
Figure 17.15 amino acid)
Building a Polypeptide
We can divide translation into three stages
Initiation
Elongation
Termination
The AUG start codon is recognized by methionyl-tRNA
or Met
Once the start codon has been identified, the ribosome
incorporates amino acids into a polypeptide chain
RNA is decoded by tRNA (transfer RNA) molecules,
which each transport specific amino acids to the
growing chain
Translation ends when a stop codon (UAA, UAG, UGA)
is reached
Initiation of Translation
The initiation stage of translation brings together
mRNA, tRNA bearing the first amino acid of the
polypeptide, and two subunits of a ribosome
Large
ribosomal
P site subunit
3 U A C 5
5 A U G 3
Initiator tRNA
GTP GDP
E A
mRNA
5 3 5 3
Start codon
1 2
A small ribosomal subunit binds to a molecule of The arrival of a large ribosomal subunit completes
mRNA. In a prokaryotic cell, the mRNA binding site the initiation complex. Proteins called initiation
on this subunit recognizes a specific nucleotide factors (not shown) are required to bring all the
sequence on the mRNA just upstream of the start translation components together. GTP provides
codon. An initiator tRNA, with the anticodon UAC, the energy for the assembly. The initiator tRNA is
base-pairs with the start codon, AUG. This tRNA in the P site; the A site is available to the tRNA
carries the amino acid methionine (Met). bearing the next amino acid.
Elongation of the Polypeptide Chain
In the elongation stage, amino acids are added one
by one to the preceding amino acid
E E
P A P A
5
3 3
3
5 5
Stop codon
(UAG, UAA, or UGA)
1 When a ribosome reaches a stop 2 The release factor hydrolyzes 3 The two ribosomal subunits
codon on mRNA, the A site of the the bond between the tRNA in and the other components of
ribosome accepts a protein called the P site and the last amino the assembly dissociate.
a release factor instead of tRNA. acid of the polypeptide chain.
The polypeptide is thus freed
from the ribosome.
Translation: Initiation
mRNA binds to a ribosome, and the transfer RNA corresponding
to the START codon binds to this complex. Ribosomes are
composed of 2 subunits (large and small), which come together
when the messenger RNA attaches during the initiation process.
Translation: Elongation
Elongation: the ribosome moves down the messenger RNA,
adding new amino acids to the growing polypeptide chain.
The ribosome has 2 sites for binding transfer RNA. The first
RNA with its attached amino acid binds to the first site, and then
the transfer RNA corresponding to the second codon bind to the
second site.
Translation: Elongation
The ribosome then removes the amino acid from the
first transfer RNA and attaches it to the second amino
acid.
At this point, the first transfer RNA is empty: no
attached amino acid, and the second transfer RNA
has a chain of 2 amino acids attached to it.
Translation: Termination
The elongation cycle repeats as the ribosome moves
down the messenger RNA, translating it one codon
and one amino acid at a time.
The process repeats until a STOP codon is reached.
Polyribosomes
A number of ribosomes can translate a single mRNA
molecule simultaneously forming a polyribosome
Polyribosomes enable a cell to make many copies of
a polypeptide very quickly
Completed
Growing polypeptide
polypeptides
Incoming
ribosomal
subunits
Start of End of
mRNA mRNA
(5 end) (3 end)
(a) An mRNA molecule is generally translated simultaneously
by several ribosomes in clusters called polyribosomes.
Ribosomes
mRNA
0.1 m
This micrograph shows a large polyribosome in a prokaryotic
cell (TEM).
Comparing Gene Expression In Prokaryotes And Eukaryotes
In a eukaryotic cell:
The nuclear envelope separates transcription from translation
Extensive RNA processing occurs in the nucleus
Prokaryotic cells lack a nuclear envelope, allowing translation to
begin while transcription progresses
RNA polymerase
DNA
mRNA
Polyribosome
Direction of 0.25 mm
RNA transcription
polymerase DNA
Polyribosome
Polypeptide
(amino end)
Ribosome
mRNA (5 end)
A summary of transcription and translation in a eukaryotic cell
TRANSCRIPTION DNA
1RNA is transcribed
from a DNA template.
3
5 RNA RNA
transcript polymerase
RNA PROCESSING Exon
2
In eukaryotes, the RNA transcript
RNA transcript (pre- (pre-mRNA)
mRNA) is spliced and Intron
modified to produce
mRNA, which moves Aminoacyl-tRNA
from the nucleus to the synthetase
cytoplasm. NUCLEUS
Amino
FORMATION OF acid
INITIATION COMPLEX AMINO ACID ACTIVATION
CYTOPLASM tRNA
3 After leaving the 4
Each amino acid
nucleus, mRNA attaches attaches to its proper tRNA
to the ribosome. with the help of a specific
enzyme and ATP.
mRNA Growing
polypeptide
Activated
amino acid
Ribosomal
subunits
5
TRANSLATION
5
A succession of tRNAs
E A add their amino acids to
Anticodon the polypeptide chain
A A A
as the mRNA is moved
U G G U U U A U G
through the ribosome
one codon at a time.
Figure 17.26 Codon (When completed, the
polypeptide is released
Ribosome from the ribosome.)
Post-translation
The new polypeptide is now floating loose in the
cytoplasm if translated by a free ribosome.
Polypeptides fold spontaneously into their active
configuration, and they spontaneously join with other
polypeptides to form the final proteins.
Often translation is not sufficient to make a functional
protein, polypeptide chains are modified after
translation
Sometimes other molecules are also attached to the
polypeptides: sugars, lipids, phosphates, etc. All of
these have special purposes for protein function.
Targeting Polypeptides to Specific Locations
Completed proteins are targeted to specific sites
in the cell
Two populations of ribosomes are evident in cells:
free ribsomes (in the cytosol) and bound
ribosomes (attached to the ER)
Free ribosomes mostly synthesize proteins that
function in the cytosol
Bound ribosomes make proteins of the endomembrane
system and proteins that are secreted from the cell
Ribosomes are identical and can switch from free
to bound
Targeting Polypeptides to Specific Locations
Polypeptide synthesis always begins in the cytosol
Synthesis finishes in the cytosol unless the polypeptide signals the ribosome to
attach to the ER
Polypeptides destined for the ER or for secretion are marked by a signal
peptide
A signal-recognition particle (SRP) binds to the signal peptide
The SRP brings the signal peptide and its ribosome to the ER
Ribosomes
mRNA
Signal
peptide ER
Signal- membrane
Signal
recognition
peptide Protein
particle SRP
removed
(SRP) receptor
protein
CYTOSOL
ER LUMEN Translocation
complex
Mutation Causes and Rate
The natural replication of DNA produces occasional
errors. DNA polymerase has an editing mechanism
that decreases the rate, but it still exists
Typically genes incur base substitutions about once
in every 10,000 to 1,000,000 cells
Since we have about 6 billion bases of DNA in each
cell, virtually every cell in your body contains several
mutations
Mutations can be harmful, lethal, helpful, silent
However, most mutations are neutral: have no effect
Only mutations in cells that become sperm or eggs
are passed on to future generations
Mutations in other body cells only cause trouble when
they cause cancer or related diseases
Mutagens
Mutagens are chemical or physical agents that interact
with DNA to cause mutations.
Physical agents include high-energy radiation like X-rays
and ultraviolet light
Chemical mutagens fall into several categories.
Chemicals that are base analogues that may be substituted into
DNA, but they pair incorrectly during DNA replication.
Interference with DNA replication by inserting into DNA and
distorting the double helix.
Chemical changes in bases that change their pairing properties.
Tests are often used as a preliminary screen of chemicals
to identify those that may cause cancer
Most carcinogens are mutagenic and most mutagens are
carcinogenic.
Viral Mutagens
Scientists have recognized a number of
tumor viruses that cause cancer in various
animals, including humans
About 15% of human cancers are caused by
viral infections that disrupt normal control of
cell division
All tumor viruses transform cells into cancer
cells through the integration of viral nucleic
acid into host cell DNA.
Point mutations
Point mutations involve alterations in the
structure or location of a single gene.
Generally, only one or a few base pairs are
involved.
Point mutations can signficantly affect protein
structure and function
Point mutations may be caused by physical
damage to the DNA from radiation or
chemicals, or may occur spontaneously
Point mutations are often caused by mutagens
Point Mutation
The change of a single nucleotide in the DNAs template
strand leads to the production of an abnormal protein
mRNA mRNA
The mutant mRNA has
G A A G U A a U instead of an A in
one codon.
5 3 5 3
A U G A A G U U U G G U U A A
A U G A A G U U U A G U U A A
A U G U A G U U U G G C U A A
Met Stop
Insertions and Deletions
Are additions or losses of nucleotide pairs in a gene
May produce frameshift mutations that will change the
reading frame of the gene, and alter all codons
downstream from the mutation.
Wild type
A U GA A GU U U G G C U A A
mRNA 5 3
Protein Met Lys Phe Gly Stop
Amino end Carboxyl end
Base-pair insertion or deletion
Frameshift causing immediate nonsense
Extra U
AU G U A AG U U U G GC U A
Met Stop
Frameshift causing
extensive missense U Missing
A U G A A GU U G G C U A A
Met Lys Leu Ala
Insertion or deletion of 3 nucleotides:
no frameshift but extra or missing amino acid
A A G Missing
A U G U U U G G C U A A
Met Phe Gly
Stop