Chapter 17

From Gene to Protein

AP Biology

Fig. 17-1

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

Concept 17.1: Genes specify proteins via

transcription and translation

Linking Genes to Enzymes

• 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

– He referred to such errors as “inborn errors of metabolism”

• Ex) Alkaptonuria results from inability to make the enzyme

alkapton

• Linking genes to enzymes required understanding that cells synthesize and

degrade molecules in a series of steps, a metabolic pathway

Fig. 17-2a

EXPERIMENT

Growth:

Wild-type

cells growing

and dividing

No growth:

Mutant cells

cannot grow

and divide

Minimal medium

Nutritional Mutants in Neurospora

• A breakthrough in the relationship between genes and enzymes came with the work

of George Beadle and Edward Tatum

– Exposed the bread mold Neurospora crassa to X-rays

– This created mutants that were unable to survive on a minimal medium (agar)

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

Fig. 17-2b

RESULTSClasses of Neurospora crassa

Wild type Class I mutants Class II mutantsClass III mutants

Minimal

medium

(MM)

(control)

MM +

ornithine

MM +

citrulline

MM +

arginine

(control)

Co

nd

itio

n

Beadle and Tatum’s Experiment

• Beadle and Tatum showed that the Neurospora crassa mutants fell into 3 classes,

each defective in a different gene

– They grew these 3 classes of mutants under 4 different conditions

• The wild-type strain was capable of growth under all experimental

conditions, requiring only minimal medium

• The 3 classes of mutants each had a specific set of growth requirements

– Ex) Class II mutants

could not grow when

ornithine alone

was added but

could grow when

either citrulline

or arginine was

added

Fig. 17-2c

CONCLUSION Class I mutants

(mutation in

gene A)

Class II mutants

(mutation in

gene B)

Class III mutants

(mutation in

gene C)Wild type

Precursor Precursor Precursor PrecursorEnzyme AEnzyme AEnzyme AEnzyme A

Ornithine Ornithine Ornithine Ornithine

Enzyme BEnzyme B Enzyme BEnzyme B

Citrulline Citrulline Citrulline Citrulline

Enzyme CEnzyme CEnzyme CEnzyme C

Arginine Arginine Arginine Arginine

Gene A

Gene B

Gene C

Beadle and Tatum’s Experiment Continued

• Based on the growth requirements of these mutants, Beadle and Tatum deduced

that each class of mutants was unable to carry out one step in the pathway for

synthesizing arginine because it lacked the necessary enzyme

– Because each mutant was mutated in only a single gene, they concluded that

each mutated gene must normally dictate the production of one enzyme

– Their results supported the one gene-one enzyme hypothesis and also

confirmed the arginine pathway

The Products of Gene Expression: A Developing Story

• As researchers learned more about proteins, they made revisions to the one gene-

one enzyme hypothesis

– Some proteins aren’t enzymes, so researchers later renamed the hypothesis to

read one gene–one protein

– Many proteins are composed of several polypeptides, each of which has its

own gene

• Ex) Hemoglobin is built from 2 polypeptides, and thus 2 genes code for this

single protein

– 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

more precisely as polypeptides

Basic Principles of Transcription and Translation

• Though genes provide the instructions for making specific proteins, they do not build

proteins directly

– 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

• The central dogma is the concept that cells are governed by a cellular chain of

command: DNA RNA protein

Fig. 17-3a-2

(a) Bacterial cell

TRANSCRIPTIONDNA

mRNA

TRANSLATIONRibosome

Polypeptide

Protein Synthesis in Prokaryotes

• Though the basic mechanics of protein synthesis are similar for bacteria and

eukaryotes, there are important differences

• Because prokaryotes lack nuclei, the lack of segregation between DNA

and ribosomes allows translation of mRNA to begin while transcription

is still in progress

• The mRNA produced by

transcription is immediately

translated without

more processing

Fig. 17-3b-3

(b) Eukaryotic cell

TRANSCRIPTION

Nuclearenvelope

DNA

Pre-mRNARNA PROCESSING

mRNA

TRANSLATION Ribosome

Polypeptide

Protein Synthesis in Eukaryotes

• In eukaryotic cells, the nuclear envelope separates transcription and translation in

space and time

• Transcription occurs in the nucleus, and the mRNA is then transported to the

cytoplasm, where translation occurs

• Eukaryotic RNA transcripts are also modified through RNA processing to yield

finished mRNA

• A primary transcript (or pre-mRNA)

is the initial RNA transcript from any

gene, including those coding for RNA

that are not translated into protein

The Genetic Code

• Biologists encountered a problem when they began to suspect that the

instructions for protein synthesis were encoded in DNA

– There are 20 amino acids, but there are only four nucleotide bases in

DNA

– Therefore, how many bases correspond to an amino acid?

• 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 (43 = 64)

• Ex) AGT = serine

Transcription of the DNA Template

• For each gene, only one of the 2 DNA strands is transcribed

– This strand is called the template strand because it provides the pattern (or

template) for the sequence of nucleotides in an RNA transcript

• A given DNA strand is the template strand for some genes, while for other

genes, the complementary strand functions as a template

– An mRNA is complementary rather than identical to its DNA template

• RNA bases are assembled according to base-pairing rules

• These pairs are similar to those that form during DNA replication, with one

exception

– The RNA substitute for thymine that pairs with adenine is called uracil

(U)

Fig. 17-4

DNAmolecule

Gene 1

Gene 2

Gene 3

DNAtemplatestrand

TRANSCRIPTION

TRANSLATION

mRNA

Protein

Codon

Amino acid

Codons

• 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

Fig. 17-5

Second mRNA base

Fir

st

mR

NA

ba

se (

5en

d o

f co

do

n)

Th

ird

mR

NA

bas

e (

3en

d o

f co

do

n)

Cracking the Code

• All 64 codons were deciphered by the mid-1960s

– 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

Fig. 17-6

(a) Tobacco plant expressing

a firefly gene

(b) Pig expressing a

jellyfish gene

Evolution of the Genetic Code

• The genetic code is nearly universal, shared by the simplest bacteria to the most

complex animals

– Ex) CCG is translated into the amino acid proline in all organisms

• In laboratory experiments, genes can be transcribed and translated after being

transplanted from one species to

another

– Ex) Bacteria can be

programmed to synthesize

human proteins (insulin) for

medical use by the insertion

of human genes

Concept Check 17.1

• 1) What polypeptide product would you expect from a poly-G mRNA that is

30 nucleotides long?

• 2) The template strand of a gene contains the sequence 3’-TTCAGTCGT-5’.

Draw the nontemplate sequence and the mRNA sequence, indicating the 5’

and 3’ ends of each. Compare the two sequences.

• 3) Imagine that the nontemplate sequence in question 2 was transcribed

instead of the template sequence. Draw the mRNA sequence and translate

it using Figure 17.5 (pp. 330). (Be sure to pay attention to the 5’ and 3’

ends.) Predict how well the protein synthesized from the nontemplate strand

would function, if at all.

Concept 17.2: Transcription is the DNA-directed

synthesis of RNA: a closer look

Transcription: A Closer Look

• RNA synthesis is catalyzed by an enzyme called RNA polymerase

– This enzyme pries the DNA strands apart and hooks together the RNA

nucleotides

• 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

– RNA synthesis follows the same base-pairing rules as DNA, except

uracil substitutes for thymine

• There are 3 stages of transcription: initiation, elongation, and termination

Fig. 17-7a-2Promoter Transcription unit

DNAStart point

RNA polymerase

553

3

Initiation

33

1

RNAtranscript

55

UnwoundDNA

Template strandof DNA

Stages of Transcription: Initiation

Step 1: Initiation

• RNA polymerase binds to the promoter

• DNA strands unwind

• Polymerase initiates RNA synthesis at the start point on the template

strand

Fig. 17-7a-3Promoter Transcription unit

DNAStart point

RNA polymerase

553

3

Initiation

33

1

RNAtranscript

55

UnwoundDNA

Template strandof DNA

2 Elongation

RewoundDNA

5

553 3 3

RNAtranscript

Stages of Transcription: Elongation

• Step 2: Elongation

– As RNA polymerase moves “downstream,” the enzyme unwinds DNA and

elongates the RNA transcript 5’3’

• Biologists refer to the direction of transcription as “downstream” and the

other direction as “upstream”

• These terms are also used to

describe positions of nucleotide

sequences within DNA and RNA

– Ex) The promoter sequence in

DNA is upstream from the

terminator

– In the wake of transcription, the DNA

strands reform a double helix

Fig. 17-7a-4Promoter Transcription unit

DNAStart point

RNA polymerase

553

3

Initiation

33

1

RNAtranscript

55

UnwoundDNA

Template strandof DNA

2 Elongation

RewoundDNA

5

553 3 3

RNAtranscript

3 Termination

5

5

533

3Completed RNA transcript

Stages of Transcription: Termination

• Step 3: Termination

– The RNA transcript is

eventually released

– RNA polymerase

detaches from the DNA

RNA Polymerase Binding and Initiation of Transcription

• Promoters signal the initiation of RNA synthesis

– Include the transcription start point (the nucleotide where RNA synthesis

actually begins) and extends several dozen nucleotides pairs upstream from

start point

– Determine which of the 2 DNA strands is used as a template

• A collection of proteins called transcription factors mediate the binding of RNA

polymerase and the initiation of transcription in eukaryotes

– In prokaryotes, RNA polymerase itself

recognizes and binds to the promoter

– 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

Elongation of the RNA Strand

• As RNA polymerase moves along the DNA, it untwists the

double helix, 10 to 20 bases at a time

– The enzyme adds nucleotides to the 3’ end of the new RNA

molecule at a rate of 40 nucleotides per second in

eukaryotes

• A gene can be transcribed simultaneously by several RNA

polymerases

– Congregation of many polymerase molecules increases the

amount of mRNA transcribed, allowing cells to make the

encoded proteins in large amounts

Termination of Transcription

• The mechanisms of termination are different in bacteria and eukaryotes

– In bacteria, the polymerase stops transcription at the end of the terminator

– In eukaryotes, the polymerase continues transcription after the pre-mRNA is

cleaved from the growing RNA chain

• RNA polymerase transcribes a sequence called the polyadenylation signal

sequence (AAUUAA) in pre-mRNA

• Proteins associated with new RNA transcript cut it free from polymerase at

a point ~10-35 nucleotides downstream from polyadenylation signal

• Polymerase continues transcribing DNA for 100s of nucleotides past the

site where pre-mRNA is released

• The polymerase eventually falls off the DNA

Concept Check 17.2

• 1) Compare DNA polymerase and RNA polymerase in terms of how they

function, the requirement for a template and primer, the direction of

synthesis, and the type of nucleotides used.

• 2) What is a promoter, and is it located at the upstream or downstream end

of a transcription unit

• 3) What makes RNA polymerase start transcribing a gene at the right place

on the DNA in a bacterial cell? In a eukaryotic cell?

• 4) Suppose X-rays caused a sequence change in the TATA box of a

particular gene’s promoter. How would that affect transcription of the gene?

(See Figure 17.8, pp. 333).

Concept 17.3: Eukaryotic cells modify RNA after

transcription

Eukaryotic cells modify RNA after transcription

• Enzymes in the eukaryotic nucleus modify pre-mRNA 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

– These modifications produce an mRNA molecule ready for

translation

Fig. 17-9

Protein-coding segment Polyadenylation signal

3

3 UTR5 UTR

5

5 Cap Start codon Stop codon Poly-A tail

G P PP AAUAAA AAA AAA…

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 consisting of a modified

guanine nucleotide

• Occurs after transcription of the first 20-40 nucleotides

– The 3 end gets a poly-A tail before mRNA exits the nucleus

• Consists of 50-250 additional adenine nucleotides following the

polyadenylation signal (AAUAAA)

• These modifications share several functions:

– They seem to facilitate the export of mRNA from nucleus

– They protect mRNA from degradation by hydrolytic enzymes

– They help ribosomes attach to the 5 end of mRNA once it reaches the

cytoplasm

Fig. 17-10

Pre-mRNA

mRNA

Codingsegment

Introns cut out andexons spliced together

5 Cap

Exon Intron5

1 30 31 104

Exon Intron

105

Exon

146

3

Poly-A tail

Poly-A tail5 Cap

5 UTR 3 UTR1 146

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

Fig. 17-11-3

RNA transcript (pre-mRNA)

Exon 1 Exon 2Intron

Protein

snRNA

snRNPs

Otherproteins

5

5

Spliceosome

Spliceosomecomponents

Cut-outintron

mRNA

Exon 1 Exon 25

snRNPs and Spliceosomes

• The signal for RNA splicing is a short nucleotide sequence at each end of an intron

– Particles called small nuclear ribonucleoproteins (snRNPs) recognize these

splice sites

• The RNA in a snRNP particle is called

a small nuclear RNA (snRNA)

– Several different snRNPs join with additional

proteins to form even larger assemblies

called spliceosomes

• Spliceosomes interact with certain sites along

an intron, releasing the intron and

joining together the 2 exons that

flanked the intron

Ribozymes

• In some organisms, RNA splicing can occur without proteins or even additional RNA

molecules

– Ribozymes are catalytic RNA molecules that function as enzymes and can

splice RNA

– Some intron RNA functions as a ribozyme and catalyze their own excision

• The discovery of ribozymes rendered obsolete the belief that all biological catalysts

were proteins

– Three properties of RNA enable it to function as an enzyme

• It can form a three-dimensional structure because of its ability to base pair

with itself, as well as its single-stranded structure

• Some bases in RNA contain functional groups that can participate in

reactions

• RNA may hydrogen-bond with other nucleic acid molecules

The Functional and Evolutionary Importance of Introns

• A direct consequence of the presence of introns in genes is that a single

gene can encode more than one kind of polypeptide

– Many genes can give rise to 2+ different polypeptides depending on

which segments are treated as exons during RNA processing

• Such variations are called alternative RNA splicing

– Ex) Sex differences in fruit flies are largely due to differences in

how males and females splice their transcribed RNA

– Because of alternative splicing, the number of different proteins an

organism can produce is much greater than its number of genes

Fig. 17-12

Gene

DNA

Exon 1 Exon 2 Exon 3Intron Intron

Transcription

RNA processing

Translation

Domain 2

Domain 3

Domain 1

Polypeptide

Domains

• Proteins often have a modular architecture consisting of discrete regions called

domains

– Ex) One domain may include the active site

while another might attach the it to the

cell membrane

• In many cases, different exons code for the different

domains in a protein

– Exon shuffling may result in evolution of

new proteins

• Introns increase the probability of crossing over between exons of

alleles by providing more terrain for crossovers without interrupting coding

sequences

• Occasional mixing and matching of exons between completely different

(nonallelic) genes may also occur

Concept Check 17.3

• 1) How does alteration of the 5’ and 3’ ends of pre-mRNA affect

the mRNA that exits the nucleus?

• 2) How is RNA splicing similar to editing a video?

• 3) In nematode worms, a gene that codes for an ATPase has

two alternatives for exon 4 and three alternatives for exon 7.

How many different forms of the protein could be made from

this gene?

Concept 17.4: Translation is the RNA-directed

synthesis of a polypeptide: a closer look

Fig. 17-13

Polypeptide

Ribosome

Aminoacids

tRNA withamino acidattached

tRNA

Anticodon

Trp

Phe Gly

Codons 35

mRNA

Transfer RNA

• The translation of mRNA to protein can be examined in more detail

– A cell translates an mRNA message into protein with the help of transfer RNA

(tRNA)

• Molecules of tRNA are not identical:

– Each carries a specific amino acid on one end

– Each has an anticodon on the other end

• The anticodon base-pairs with a

complementary codon on mRNA

Fig. 17-14

Amino acidattachment site

3

5

Hydrogenbonds

Anticodon

(a) Two-dimensional structure

Amino acidattachment site

5

3

Hydrogenbonds

3 5

AnticodonAnticodon

(c) Symbol used

in this book(b) Three-dimensional structure

The Structure and Function of Transfer RNA

A C C

• A tRNA molecule consists of a single RNA strand that is only about 80

nucleotides long

– Flattened into one plane to reveal its base pairing,

a tRNA molecule looks like a cloverleaf

– Because of hydrogen bonds, tRNA actually twists

and folds into a three-dimensional molecule

• tRNA is roughly L-shaped

Steps Required for Accurate Translation

• Accurate translation requires two steps:

– First: a correct match between a tRNA and an amino acid, done by the enzyme

aminoacyl-tRNA synthetase

– Second: a correct match between the tRNA anticodon and an mRNA codon

• Some tRNAs can bond to more than one codon because the rules for

base-pairing between the 3rd base of a codon and its corresponding

anticodon are more relaxed than those of the 1st two bases

– Ex) Uracil at the 5’ end of a tRNA anticodon can pair with either

adenine or guanine in the 3rd position of an mRNA codon

– This flexible pairing at the third base of a codon is called wobble

• Wobble explains why codon synonymous for an amino acid can

differ in their 3rd base

Fig. 17-15-4

Amino acid Aminoacyl-tRNAsynthetase (enzyme)

ATP

AdenosineP P P

AdenosineP

PP i

PPi

i

tRNA

tRNA

Aminoacyl-tRNAsynthetase

Computer model

AMPAdenosineP

Aminoacyl-tRNA(“charged tRNA”)

Matching tRNA and Amino Acids

• Aminoacyl-tRNA sythetases catalyze the covalent attachment of amino

acids to their appropriate tRNA in a process driven by the hydrolysis of ATP

– 1) The active site binds the amino acid

and ATP

– 2) ATP loses 2 phosphate groups and

joins the amino acid as AMP

– 3)The appropriate tRNA covalently binds to

the amino acid, displacing AMP

– 4)The tRNA and its bound amino acid is

released by the enzyme

Fig. 17-16Growingpolypeptide Exit tunnel

Largesubunit

Smallsubunit

tRNAmolecules

E PA

mRNA5 3

(a) Computer model of functioning ribosome

P site (Peptidyl-tRNAbinding site)

E site(Exit site)

A site (Aminoacyl-tRNA binding site)

E P A Largesubunit

mRNAbinding site

Smallsubunit

(b) Schematic model showing binding sites

Amino end Growing polypeptide

Next amino acidto be added topolypeptide chain

mRNAtRNAE

3

5 Codons

(c) Schematic model with mRNA and tRNA

Ribosomes

• Ribosomes facilitate specific coupling of tRNA anticodons with mRNA codons in

protein synthesis

– Ribosomes are made up of 2 subunits, called the

large and small subunits

• These two ribosomal subunits are made of

proteins and ribosomal RNA (rRNA)

– A ribosome has three binding sites for tRNA:

• The P site holds the tRNA that carries the carries

growing polypeptide chain

• The A site holds the tRNA that carries the

next amino acid to be added to the chain

• The E site is the exit site, where discharged

tRNAs leave the ribosome

Building a Polypeptide

• The three stages of translation:

– Initiation

– Elongation

– Termination

• All three stages require protein “factors” that aid in the translation process

– Energy is also required for certain aspects of chain initiation and

elongation

• This energy is provided by the hydrolysis of GTP

Fig. 17-17

3

35

5U

U

AA

C

GMet

GTP GDPInitiator

tRNA

mRNA

53

Start codon

mRNA binding site

Smallribosomalsubunit

5

P site

Translation initiation complex

3

E A

Met

Largeribosomalsubunit

Ribosome Association and Initiation of Translation

• The initiation stage of translation brings together mRNA, a tRNA with the first amino

acid, and the two ribosomal subunits

– First, a small ribosomal subunit binds with mRNA and a special initiator tRNA

carrying the amino acid methionine

– Then the small subunit moves along the mRNA until it reaches the start codon

(AUG)

– Proteins called initiation factors bring in the large subunit that completes the

translation initiation complex

• The cell must expend energy in the form of a GTP molecule to form this

initiation complex

– At the completion of the initiation process, the initiator tRNA sits in the P site of

the ribosome

• The A site is vacant and ready

for the next aminoacyl

tRNA

Elongation of the Polypeptide Chain

• During the elongation stage, amino acids are added one by one to the

preceding amino acid

– Each addition involves several proteins called elongation factors and

occurs in three steps

• 1) Codon recognition

• 2) Peptide bond formation

• 3) Translocation

– Two molecules are GTP are required, one for codon recognition and

the other for translocation

Fig. 17-18-2

Amino endof polypeptide

mRNA

5

3E

Psite

Asite

GTP

GDP

E

P A

Steps in the Elongation of a Polypeptide Chain

• Step 1 - Codon recognition: the anticodon of an incoming aminoacyl tRNA base-

pairs with the complementary mRNA codon in the A site

– Hydrolysis of GTP increases the accuracy and efficiency of this step

• Step 2 - Peptide bond formation: an rRNA molecule of the large ribosomal subunit

catalyzes the formation of a peptide bond between the new amino acid in the A site

and the carboxyl end of the growing polypeptide in the P site

– This step removes the polypeptide from

the tRNA in the P site and attaches

it to the amino acid on the tRNA

in the A site

Fig. 17-18-4

Amino endof polypeptide

mRNA

5

3E

Psite

Asite

GTP

GDP

E

P A

E

P A

GDP

GTP

Ribosome ready fornext aminoacyl tRNA

E

P A

Steps in the Elongation of a Polypeptide Chain

• Step 3 – Translocation: the ribosome translocates the tRNA in the A site to the P

site

– At the same time, the empty tRNA in the P site is moved to the E site, where it

is released

– The mRNA then moves along with

its bound tRNAs, bringing the next

codon to be translated into

the A site

Fig. 17-19-3

Releasefactor

3

5

Stop codon(UAG, UAA, or UGA)

5

3

2

Freepolypeptide

2 GDP

GTP

5

3

Termination of Translation • Termination occurs when a stop codon in the mRNA reaches the A site of the

ribosome

– The base triplets UAG, UAA, and UGA act as signals to stop translation

– A protein called a release factor binds directly to the stop codon in the A site

• The release factor causes the addition of a water molecule instead of an

amino acid

• This reaction breaks (hydrolyzes) the bond between the completed polypeptide and

the tRNA in the P site

– This releases the polypeptide through the exit tunnel of the ribosome’s large

subunit

– The remainder of the translation assembly then comes apart in a multistep

process, aided by other protein factors

• This breakdown requires the hydrolysis of 2 more GTP molecules

Animation: Translation

Fig. 17-20

Growingpolypeptides

Completedpolypeptide

Incomingribosomalsubunits

Start ofmRNA(5 end)

Polyribosome

End ofmRNA(3 end)

(a)

Ribosomes

mRNA

(b) 0.1 µm

Polyribosomes

• A number of ribosomes can translate a single mRNA simultaneously, forming a

polyribosome (or polysome)

– Once a ribosome moves past the start

codon, a second ribosome can attach

to the mRNA

• Polyribosomes enable a cell to make

many copies of a polypeptide very

quickly

Completing and Targeting the Functional Protein

• Often translation is not sufficient to make a

functional protein

– Polypeptide chains are modified after

translation

– Completed proteins are then targeted to

specific sites in the cell

Protein Folding and Post-Translational Modifications

• 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

– Certain amino acids may be chemically modified by the attachment of sugars,

lipids, phosphate groups, or other additions

– Enzymes may remove one or more amino acids from the leading (amino) end

of the polypeptide chain

– A polypeptide chain may also be cleaved by enzymes into two or more pieces

(insulin)

• In other cases, two or more polypeptides that are synthesized separately

may come together, becoming subunits of a protein with a quaternary

structure (hemoglobin)

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

Targeting Polypeptides to Specific Locations

• What determines whether a ribosome will be free in the cytosol or bound to rough ER

at any given time?

• Polypeptide synthesis always begins in the cytosol

• The process continues there until completion unless the growing polypeptide

itself cues the ribosome to attach to the ER

• Polypeptides destined for the ER or for secretion are marked by a signal

peptide that targets the protein to the ER

• The signal peptide is a sequence of ~20 amino acids at or near the

leading (amino) end of the polypeptide

Fig. 17-21

Ribosome

mRNA

Signalpeptide

Signal-recognitionparticle (SRP)

CYTOSOLTranslocationcomplex

SRPreceptorprotein

ER LUMEN

Signalpeptideremoved

ERmembrane

Protein

Fig. 17-21

Ribosome

mRNA

Signalpeptide

Signal-recognitionparticle (SRP)

CYTOSOLTranslocationcomplex

SRPreceptorprotein

ER LUMEN

Signalpeptideremoved

ERmembrane

Protein

SRPs • The signal peptide is recognized as it emerges from the ribosome by a protein-RNA

complex called a signal-recognition particle (SRP)

• The SRP brings the signal peptide and its ribosome to a receptor protein built

into the ER membrane

• Polypeptide synthesis continues here, and the growing polypeptide moves

into the ER lumen via a protein pore

• The signal peptide is usually removed by an enzyme

• If it is to be secreted from the cell, the rest of the completed polypeptide in

released into solution in within the ER lumen

• If the polypeptide is to be a membrane protein, it remains partially

embedded in the ER membrane

Concept Check 17.4

• 1) What two processes ensure that the correct amino acid is

added to a growing polypeptide chain?

• 2) Describe how the formation of polyribosomes can benefit the

cell.

• 3) Describe how a polypeptide to be secreted is transported to

the endomembrane system.

• 4) Discuss the ways in which rRNA structure likely contributes

to ribosomal function.

Concept 17.5: Point mutations can affect protein

structure and function

Fig. 17-22

Wild-type hemoglobin DNA

mRNA

Mutant hemoglobin DNA

mRNA

3

3

3

3

3

3

5

5

5

5

5

5

C CT T T

TG GA A A

A

A A AGG U

Normal hemoglobin Sickle-cell hemoglobin

Glu Val

Point Mutations

• 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

• Ex) The genetic basis of sickle cell disease is the mutation of a single base

pair in the gene that encodes one of the polypeptides of hemoglobin

Types of Point Mutations

• Point mutations within a gene can be divided

into two general categories

– Base-pair substitutions

– Base-pair insertions or deletions

Substitutions

• A base-pair substitution replaces one nucleotide and its partner with

another pair of nucleotides

– Some substitutions are called silent mutations

• These substitutions have no effect on the amino acid produced by

a codon because of redundancy in the genetic code

– Missense mutations still code for an amino acid, but not necessarily

the right amino acid

– Nonsense mutations change an amino acid codon into a stop codon,

nearly always leading to a nonfunctional protein

Fig. 17-23a

Wild type

3 DNA template strand

3

3 5

5

5 mRNA

Protein

Amino end

Stop

Carboxyl end

A instead of G

3

3

3

U instead of C

5

5

5

Stop

Silent (no effect on amino acid sequence)

Fig. 17-23b

Wild type

DNA template strand

3

5

mRNA

Protein

5

Amino end

Stop

Carboxyl end

5

3

3

T instead of C

A instead of G

3

3

3

5

5

5

Stop

Missense

Fig. 17-23c

Wild type

DNA template strand

3

5

mRNA

Protein

5

Amino end

Stop

Carboxyl end

5

3

3

A instead of T

U instead of A

3

3

3

5

5

5

Stop

Nonsense

Insertions and Deletions

• Insertions and deletions are additions or losses of nucleotide pairs in a

gene

– These mutations have a disastrous effect on the resulting protein more

often than substitutions do

– Insertion or deletion of nucleotides may alter the reading frame,

producing a frameshift mutation

• All the nucleotides that are downstream of the deletion or insertion

will be improperly grouped into codons

• Unless the frameshift is very near the end of a gene, the protein is

almost certain to be nonfunctional

Fig. 17-23d

Wild type

DNA template strand

3

5

mRNA

Protein

5

Amino end

Stop

Carboxyl end

5

3

3

Extra A

Extra U

3

3

3

5

5

5

Stop

Frameshift causing immediate nonsense (1 base-pair insertion)

Fig. 17-23e

Wild type

DNA template strand

3

5

mRNA

Protein

5

Amino end

Stop

Carboxyl end

5

3

3

missing

missing

3

3

3

5

5

5

Frameshift causing extensive missense (1 base-pair deletion)

Fig. 17-23f

Wild type

DNA template strand

3

5

mRNA

Protein

5

Amino end

Stop

Carboxyl end

5

3

3

missing

missing

3

3

3

5

5

5

No frameshift, but one amino acid missing (3 base-pair deletion)

Stop

Mutagens

• Mutations can arise in a number of ways

– Spontaneous mutations can occur during DNA replication,

recombination, or repair

• In both prokaryotes and eukaryotes, the rate of spontaneous

mutation is about one in every 1010 nucleotide

– A number of physical and chemical agents (mutagens) can also

interact with DNA in ways that cause mutations

• Ex) X-rays and other forms of high-energy radiation are physical

mutagens

Concept Check 17.5

• 1) What happens when one nucleotide pair is lost from the

middle of the coding sequence of a gene?

• 2) A gene whose template strand contains the sequence 3’-

TACTTGTCCGATATC-5’ is mutated to 3’-

TACTTGTCCAATATC-5’. For both normal and mutant genes,

draw the double-stranded DNA, the resulting mRNA, and the

amino acid sequence each encodes. What is the effect of the

mutation on the amino acid sequence?

Concept 17.6: While gene expression differs among the domains of life, the concept of a

gene is universal

Fig. 17-24

RNA polymerase

DNA

Polyribosome

mRNA

0.25 µmDirection oftranscription

DNA

RNApolymerase

Polyribosome

Polypeptide(amino end)

Ribosome

mRNA (5 end)

Gene Expression in Prokaryotes vs. Eukaryotes

• Prokaryotes in domain Archaea share many features of gene expression with

eukaryotes

– Bacteria and eukarya differ in their RNA polymerases, termination of

transcription and ribosomes, while 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

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

• In summary, a gene can be defined as a region of DNA that can be

expressed to produce a final functional product, either a polypeptide or an

RNA molecule

Concept Check 17.6

• 1) Would the coupling of processes shown in Figure

17.24 (pp. 347) be found in a eukaryotic cell? Explain.

• 2) In eukaryotic cells, mRNAs have been found to

have a circular arrangement in which the poly-A tail is

held by proteins near the 5’ end cap. How might this

increase translation efficiency?

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

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