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Chapter 17
From Gene to Protein
PowerPoint® Lecture Presentations for
Biology
Eighth Edition
Neil Campbell and Jane Reece
Lectures by Chris Romero, updated by Erin Barley with contributions from Joan Sharp
Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings
Overview: The Flow of Genetic Information
• The information content of DNA is in the form
of specific sequences of nucleotides
– The DNA inherited by an organism leads to
specific traits by dictating the synthesis of
proteins
• Proteins are the links between genotype and
phenotype
• Gene expression, the process by which DNA
directs protein synthesis, includes two stages:
transcription and translation
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Experiments Leading to Understanding the
Link between Genes and Proteins
• Nirenberg & Matthaei – determining the
language of the genetic code.
– http://bcs.whfreeman.com/thelifewire/content
/chp12/1202002.html
• Beadle & Tatum – One Gene…One Enzyme
hypothesis.
– http://www.dnalc.org/view/16360-Animation16-One-gene-makes-one-protein-.html
Evidence from the Study of Metabolic Defects
• In 1909, British physician Archibald Garrod first
suggested that genes dictate phenotypes
through enzymes that catalyze specific
chemical reactions
• He thought symptoms of an inherited disease
reflect an inability to synthesize a certain
enzyme
• Linking genes to enzymes required
understanding that cells synthesize and
degrade molecules in a series of steps, a
metabolic pathway
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Nutritional Mutants in Neurospora: Scientific
Inquiry
• George Beadle and Edward Tatum exposed
bread mold to X-rays, creating mutants that
were unable to survive on minimal medium as
a result of inability to synthesize certain
molecules
• Using crosses, they identified three classes of
arginine-deficient mutants, each lacking a
different enzyme necessary for synthesizing
arginine
• They developed a one gene–one enzyme
hypothesis, which states that each gene
dictates production of a specific enzyme
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-2
EXPERIMENT
No growth:
Mutant cells
cannot grow
and divide
Growth:
Wild-type
cells growing
and dividing
Minimal medium
RESULTS
Classes of Neurospora crassa
Wild type
Class I mutants Class II mutants Class III mutants
Condition
Minimal
medium
(MM)
(control)
MM +
ornithine
MM +
citrulline
MM +
arginine
(control)
CONCLUSION
Wild type
Precursor
Gene A
Gene B
Gene C
Class I mutants Class II mutants Class III mutants
(mutation in
(mutation in
(mutation in
gene B)
gene A)
gene C)
Precursor
Precursor
Precursor
Enzyme A
Enzyme A
Enzyme A
Enzyme A
Ornithine
Ornithine
Ornithine
Ornithine
Enzyme B
Enzyme B
Enzyme B
Enzyme B
Citrulline
Citrulline
Citrulline
Citrulline
Enzyme C
Enzyme C
Enzyme C
Enzyme C
Arginine
Arginine
Arginine
Arginine
The Products of Gene Expression: A Developing
Story
• Some proteins aren’t enzymes, so researchers
later revised the hypothesis: one gene–one
protein
• Many proteins are composed of several
polypeptides, each of which has its own gene
• Therefore, Beadle and Tatum’s hypothesis is
now restated as the one gene–one polypeptide
hypothesis
• Note that it is common to refer to gene
products as proteins rather than polypeptides
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Basic Principles of Transcription and Translation
• RNA is the intermediate between genes and the proteins for
which they code
• Transcription is the synthesis of RNA under the direction of
DNA
–
Transcription produces messenger RNA (mRNA)
• Translation is the synthesis of a polypeptide, which occurs
under the direction of mRNA
–
Ribosomes are the sites of translation
• A primary transcript is the initial RNA transcript from any gene
•
The central dogma is the concept that cells are governed by a
cellular chain of command: DNA RNA protein
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Overview of Gene Expression
• Genes contain the blueprints for building proteins
in cells.
• RNA is the bridge between DNA and its protein.
• The “genetic code” is the sequence of bases in
DNA that will code for specific amino acids in a
growing polypeptide (protein).
– There are 20 possible amino acids.
Types of RNA
•
•
RNA is a nitrogenous base that consists of a ribose (5
carbon sugar).
–
In RNA, the base Uracil is substituted for Thymine in DNA.
–
The RNA nucleic acid is single stranded.
In most cells, RNA molecules are involved in just one job
– protein synthesis!
–
•
The assembly of amino acids into proteins is controlled by RNA
There are 3 main types of RNA:
1.
Messenger RNA (mRNA)
2.
Ribosomal RNA (rRNA)
3.
Transfer RNA (tRNA)
Protein Synthesis
• Building a protein involves 2 stages:
– (1) Transcription; (2) Translation
• Transcription occurs in the nucleus
– DNA  mRNA
• Making mRNA from a DNA template in the nucleus.
• Translation occurs in the cytoplasm at a ribosome
– mRNA  protein (using tRNA and rRNA)
• Translating instructions in mRNA into a growing
polypeptide protein chain at the ribosome.
Prokaryotic v. Eukaryotic Gene Expression
http://highered.mcgraw-hill.com/olc/dl/120077/bio25.swf
The Genetic Code
• How are the instructions for assembling amino
acids into proteins encoded into DNA?
• There are 20 amino acids, but there are only
four nucleotide bases in DNA
• How many bases correspond to an amino
acid?
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Codons: Triplets of Bases
• The flow of information from gene to protein is
based on a triplet code: a series of
nonoverlapping, three-nucleotide words
• These triplets are the smallest units of uniform
length that can code for all the amino acids
• Example: AGT at a particular position on a
DNA strand results in the placement of the
amino acid serine at the corresponding position
of the polypeptide to be produced
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Codons: Triplets of Bases
• During transcription, one of the two DNA strands called the
template strand provides a template for ordering the
sequence of nucleotides in an RNA transcript
• During translation, the mRNA base triplets, called codons,
are read in the 5 to 3 direction
• Each codon specifies the amino acid to be placed at the
corresponding position along a polypeptide
• Codons along an mRNA molecule are read by translation
machinery in the 5 to 3 direction
• Each codon specifies the addition of one of 20 amino acids
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-4
DNA
molecule
Gene 2
Gene 1
Gene 3
DNA
template
strand
TRANSCRIPTION
mRNA
Codon
TRANSLATION
Protein
Amino acid
Cracking the Code
• All 64 codons were deciphered by the mid1960s
• Of the 64 triplets, 61 code for amino acids; 3
triplets are “stop” signals to end translation
• The genetic code is redundant but not
ambiguous; no codon specifies more than one
amino acid
• Codons must be read in the correct reading
frame (correct groupings) in order for the
specified polypeptide to be produced
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Third mRNA base (3 end of codon)
First mRNA base (5 end of codon)
Fig. 17-5
Second mRNA base
Fig. 17-6
(a) Tobacco plant expressing
a firefly gene
(b) Pig expressing a
jellyfish gene
Types of RNA
http://www-class.unl.edu/biochem/gp2/m_biology/animation/gene/gene_a1.html
rR
NA
Transcription
Section 12-3 http://bcs.whfreeman.com/thelifewire/content/chp12/1202001.html
Adenine (DNA and RNA)
Cystosine (DNA and RNA)
Guanine(DNA and RNA)
Thymine (DNA only)
Uracil (RNA only)
RNA
polymerase
DNA
RNA
During transcription, RNA polymerase uses one strand of DNA as a
template to assemble nucleotides into a strand of RNA.
Transcription
• Transcription, the first stage of gene expression, is the
DNA-directed synthesis of mRNA
• RNA synthesis is catalyzed by RNA polymerase, which
pries the DNA strands apart and hooks together the RNA
nucleotides
• RNA synthesis follows the same base-pairing rules as
DNA, except uracil substitutes for thymine
• The DNA sequence where RNA polymerase attaches is
called the promoter; in bacteria, the sequence signaling
the end of transcription is called the terminator
• The stretch of DNA that is transcribed is called a
transcription unit
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-7
Promoter
Transcription unit
5
3
Start point
RNA polymerase
3
5
DNA
1 Initiation
5
3
RNA
transcript
RNA
polymerase
Template strand
of DNA
3
2 Elongation
Rewound
DNA
5
3
RNA nucleotides
3
5
Unwound
DNA
3
5
5
5
Direction of
transcription
(“downstream”)
3 Termination
3
5
5
3
5
3 end
5
3
RNA
transcript
Nontemplate
strand of DNA
Elongation
Completed RNA transcript
3
Newly made
RNA
Template
strand of DNA
RNA Polymerase Binding and Initiation of
Transcription
• Promoters signal the initiation of RNA
synthesis
• Transcription factors mediate the binding of
RNA polymerase and the initiation of
transcription
• The completed assembly of transcription
factors and RNA polymerase II bound to a
promoter is called a transcription initiation
complex
• A promoter called a TATA box is crucial in
forming the initiation complex in eukaryotes
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-8
1
Promoter
A eukaryotic promoter
includes a TATA box
Template
5
3
3
5
TATA box
Start point Template
DNA strand
2
Transcription
factors
Several transcription factors must
bind to the DNA before RNA
polymerase II can do so.
5
3
3
5
3
Additional transcription factors bind to
the DNA along with RNA polymerase II,
forming the transcription initiation complex.
RNA polymerase II
Transcription factors
5
3
3
5
5
RNA transcript
Transcription initiation complex
http://bcs.whfreeman.com/thelif
ewire/content/chp14/1402002.ht
ml
Concept 17.3: Eukaryotic cells modify RNA after
transcription
• Enzymes in the eukaryotic nucleus modify premRNA before the genetic messages are
dispatched to the cytoplasm
• During RNA processing, both ends of the
primary transcript are usually altered
• Also, usually some interior parts of the
molecule are cut out, and the other parts
spliced together
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Alteration of mRNA Ends
• Each end of a pre-mRNA molecule is modified
in a particular way:
– The 5 end receives a modified nucleotide 5
cap
– The 3 end gets a poly-A tail
• These modifications share several functions:
– They seem to facilitate the export of mRNA
– They protect mRNA from hydrolytic enzymes
– They help ribosomes attach to the 5 end
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-9
50 to 250 adenine nucleotides
added to the 3’ end created by
cleavage downstream of the
termination signal.
A modified guanosine
triphosphate added to
the 5’ end.
5
G
Protein-coding segment Polyadenylation signal
3
P P P
5 Cap
AAUAAA
5 UTR Start codon
Stop codon
3 UTR
AAA…AAA
Poly-A tail
Split Genes and RNA Splicing
• Most eukaryotic genes and their RNA
transcripts have long noncoding stretches of
nucleotides that lie between coding regions
• These noncoding regions are called intervening
sequences, or introns
• The other regions are called exons because
they are eventually expressed, usually
translated into amino acid sequences
• RNA splicing removes introns and joins
exons, creating an mRNA molecule with a
continuous coding sequence
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-10
5 Exon Intron
Exon
Exon
Intron
3
Pre-mRNA 5 Cap
Poly-A tail
1
30
31
Coding
segment
mRNA 5 Cap
1
5 UTR
104
105
146
Introns cut out and
exons spliced together
Poly-A tail
146
3 UTR
Pre-RNA Splicing
http://bcs.whfreeman.com/thelifewire/content/chp14/1402001.html
• In some cases, RNA splicing is carried out by spliceosomes
• Spliceosomes consist of a variety of proteins and several
small nuclear ribonucleoproteins (snRNPs) that recognize
the splice sites
• snRNP’s : small nuclear ribonucleoproteins
–
these look for special sequences at ends of introns to know where
to cut
–
made up of snRNA (small nuclear RNA)
• Several different snRNP’s join with additional proteins to
form the splicesome – this interacts with the splice sites at
intron ends, and cuts them out – then joins the two exons
together.
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Figure 17.10 The roles of snRNPs and spliceosomes in mRNA splicing
1. Pre-mRNA
containing exons
and introns
combines with
small nuclear
ribonuceloproteins
(snRNPs) and
other proteins to
form a molecular
complex call
spliceosome.
2. Within the
spliceosome,
snRNA base-pairs
with nucleotides at
the ends of the
intron.
3. The RNA
transcript is cut to
release the intron,
and the exons are
spliced together.
The spliceosome
then comes apart,
releasing mRNA,
which now contains
only exons.
The Functional and Evolutionary Importance of
Introns
• Some genes can encode more than one kind of
polypeptide, depending on which segments are
treated as exons during RNA splicing
• Such variations are called alternative RNA
splicing
• Because of alternative splicing, the number of
different proteins an organism can produce is
much greater than its number of genes
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Translation
• Translation is the RNA-directed synthesis of a polypeptide
• Generally: is the reading of the codons on the mRNA strand
and the sequencing of them into an amino acid sequence –
polypeptide.
• The Players:
–
mRNA: already processed within the nucleus via transcription, will
be the template for the sequence of amino acids.
–
tRNA: transfers amino acids from the cytoplasm to the ribosome.
–
rRNA (ribosome): adds amino acids together from the tRNA and in
the sequence of the mRNA.
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-13
http://highered.mcgrawhill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::
535::/sites/dl/free/0072437316/120077/micro0
6.swf::Protein%20Synthesis
Amino
acids
Polypeptide
tRNA with
amino acid
attached
Ribosome
tRNA
Anticodon
Codons
5
mRNA
3
Fig. 17-14
3
Amino acid
attachment site
5
Hydrogen
bonds
Anticodon
(a) Two-dimensional structure
Amino acid
attachment site
5
3
Hydrogen
bonds
3
Anticodon
(b) Three-dimensional structure
5
Anticodon
(c) Symbol used
in this book
Fig. 17-15-4
Aminoacyl-tRNA
synthetase (enzyme)
Amino acid
P P P
Adenosine
ATP
P
P Pi
Pi
Adenosine
tRNA
Aminoacyl-tRNA
synthetase
Pi
tRNA
P
Adenosine
AMP
Computer model
Aminoacyl-tRNA
(“charged tRNA”)
Fig. 17-16
Growing
polypeptide
Exit tunnel
tRNA
molecules
EP
Large
subunit
A
Small
subunit
5
mRNA
3
(a) Computer model of functioning ribosome
P site (Peptidyl-tRNA
binding site)
E site
(Exit site)
A site (AminoacyltRNA binding site)
E P A
mRNA
binding site
Large
subunit
Small
subunit
(b) Schematic model showing binding sites
Growing polypeptide
Amino end
Next amino acid
to be added to
polypeptide chain
E
mRNA
5
tRNA
3
Codons
(c) Schematic model with mRNA and tRNA
http://highered.mcgrawhill.com/olc/dl/120077/mi
cro06.swf
Building a Polypeptide
• The three stages of translation:
– Initiation (requires energy “GTP”)
– Elongation (requires energy “GTP”)
– Termination
• All three stages require protein “factors” that
aid in the translation process
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-17
INITIATION OF TRANSLATION
3 U A C 5
5 A U G 3
Initiator
tRNA
Large
ribosomal
subunit
P site
GTP GDP
E
mRNA
5
Start codon
mRNA binding site
3
Small
ribosomal
subunit
5
A
3
Translation initiation complex
Fig. 17-18-4
ELONGATION
OF
TRANSLATION
Amino end
of polypeptide
E
3
mRNA
Ribosome ready for
next aminoacyl tRNA
P A
site site
5
GTP
GDP
E
E
P A
P A
GDP
GTP
E
P A
Fig. 17-19-3
TERMINATION OF TRANSLATION
Release
factor
Free
polypeptide
5
3
5
5
Stop codon
(UAG, UAA, or UGA)
3
2 GTP
2 GDP
3
Fig. 17-20
Growing
polypeptides
Completed
polypeptide
Incoming
ribosomal
subunits
Start of
mRNA
(5 end)
(a)
End of
mRNA
(3 end)
Ribosomes
mRNA
(b)
0.1 µm
Completing and Targeting the Functional Protein
• Often translation is not sufficient to make a functional
protein…THEREFORE…polypeptide chains are modified
after translation
• Completed proteins are targeted to specific sites in the cell
–
During and after synthesis, a polypeptide chain spontaneously
coils and folds into its three-dimensional shape
–
Proteins may also require post-translational modifications before
doing their job
–
Some polypeptides are activated by enzymes that cleave them
–
Other polypeptides come together to form the subunits of a
protein
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Targeting Polypeptides to Specific Locations
• 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
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Review – PROTEIN SYNTHESIS
• Transcription
– DNA  mRNA (in nucleus)
– In eukaryotes will have RNA processing (in nucleus).
• Translation
– mRNA  Polypeptide (at ribosome in cytoplasm)
• Coiling & Folding
– Three dimensional (in cytoplasm as translation is
occurring)
– Chaperone proteins involved
Protein Synthesis
Nucleus
Messenger RNA
Messenger RNA is transcribed in the nucleus.
Phenylalanine
tRNA
The mRNA then enters the cytoplasm and
attaches to a ribosome. Translation begins at
AUG, the start codon. Each transfer RNA has
an anticodon whose bases are complementary
to a codon on the mRNA strand. The ribosome
positions the start codon to attract its
anticodon, which is part of the tRNA that binds
methionine. The ribosome also binds the next
codon and its anticodon.
Ribosome
Go to
Section:
mRNA
Transfer RNA
Methionine
mRNA
Lysine
Start codon
Protein Synthesis Continued
Concept 17.5: Point mutations can affect protein
structure and function
• Mutations are changes in the genetic material
of a cell or virus
• Point mutations are chemical changes in just
one base pair of a gene
• The change of a single nucleotide in a DNA
template strand can lead to the production of
an abnormal protein
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-22
Wild-type hemoglobin DNA
Mutant hemoglobin DNA
C T T
C A T
3
5 3
G T A
5
G A A
3 5
mRNA
5
5
3
mRNA
G A A
Normal hemoglobin
Glu
3 5
G U A
Sickle-cell hemoglobin
Val
3
Types of Point Mutations
• Point mutations within a gene can be divided
into two general categories
– Base-pair substitutions
– Base-pair insertions or deletions
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-23
http://nortonbooks.com/col
lege/biology/animations/ch
13a08.htm
Wild-type
DNA template strand 3
5
5
3
mRNA 5
3
Protein
Stop
Amino end
Carboxyl end
A instead of G
3
5
Extra A
5
3
3
5
3
5
U instead of C
5
5
3
Extra U
3
Stop
Stop
Silent (no effect on amino acid sequence)
Frameshift causing immediate nonsense (1 base-pair insertion)
T instead of C
missing
3
5
5
3
3
5
3
5
5
3
A instead of G
missing
5
3
Stop
Missense
Frameshift causing extensive missense (1 base-pair deletion)
missing
A instead of T
5
3
3
5
U instead of A
5
5
3
3
5
missing
3
5
Stop
Stop
Nonsense
(a) Base-pair substitution
3
No frameshift, but one amino acid missing (3 base-pair deletion)
(b) Base-pair insertion or deletion
Mutagens
• Spontaneous mutations can occur during DNA
replication, recombination, or repair
• Mutagens are physical or chemical agents that
can cause mutations
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Comparing Gene Expression in Bacteria, Archaea,
and Eukarya
• Archaea are prokaryotes, but share many features of gene
expression with eukaryotes
• Bacteria and eukarya differ in their RNA polymerases,
termination of transcription and ribosomes; archaea tend to
resemble eukarya in these respects
• Bacteria can simultaneously transcribe and translate the
same gene
• In eukarya, transcription and translation are separated by
the nuclear envelope
• In archaea, transcription and translation are likely coupled
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
What Is a Gene? Revisiting the Question
• The idea of the gene itself is a unifying concept
of life
• We have considered a gene as:
– A discrete unit of inheritance
– A region of specific nucleotide sequence in a
chromosome
– A DNA sequence that codes for a specific
polypeptide chain
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
Fig. 17-25
DNA
TRANSCRIPTION
3
RNA
polymerase
5 RNA
transcript
RNA PROCESSING
Exon
RNA transcript
(pre-mRNA)
Intron
Aminoacyl-tRNA
synthetase
NUCLEUS
Amino
acid
CYTOPLASM
AMINO ACID ACTIVATION
tRNA
mRNA
Growing
polypeptide
3
A
Activated
amino acid
P
E
Ribosomal
subunits
5
TRANSLATION
E
A
Codon
Ribosome
Anticodon
The Essay – IT’S A DOOZY!
•
A portion of a specific DNA molecule consists of the
following sequence of nucleotide triplets:
TAC
•
GAA
CTT
CGG
TCC
This DNA sequence codes for the following short
polypeptide:
methionine - leucine - glutamic acid - proline - arginine
a) Describe the steps in the synthesis of this polypeptide.
b) What would be the effect of a deletion or an addition in one of the DNA
nucleotides?
c) What would be the effect of a substitution in one of the nucleotides?
d) Cells regulate both protein synthesis and protein activity. Discuss TWO
specific mechanisms of protein regulation in eukaryotic cells.
You should now be able to:
1. Describe the contributions made by Garrod,
Beadle, and Tatum to our understanding of
the relationship between genes and enzymes
2. Briefly explain how information flows from
gene to protein
3. Compare transcription and translation in
bacteria and eukaryotes
4. Explain what it means to say that the genetic
code is redundant and unambiguous
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings
5. Include the following terms in a description of
transcription: mRNA, RNA polymerase, the
promoter, the terminator, the transcription
unit, initiation, elongation, termination, and
introns
6. Include the following terms in a description of
translation: tRNA, wobble, ribosomes,
initiation, elongation, and termination
Copyright © 2008 Pearson Education Inc., publishing as Pearson Benjamin Cummings