Chapter 17Chapter 17

From Gene to Protein

DNA

• DNA structure = double helix

• DNA sugar = deoxyribose

• DNA bases = A, T, C, & G

• A T C T C G A G T C G A T T A G A G C T C A G C T A

RNA• RNA structure = usually single stranded

• RNA sugar = ribose

• RNA bases = A, U (uracil), C, & G (no thymine)

• DNA sequence: A T C T C G A G T C G A T RNA sequence: U A G A G C U C A GC U A

The Connection Between Genes & Proteins

• gene – a short sequence of DNA that codes for a specific piece of information

The Connection Between Genes & Proteins

• Beadle and Tatum

– proposed the one gene – one enzyme hypothesis

• one gene – one enzyme hypothesis – states that the function of a gene is to produce a specific enzyme

The Connection Between Genes & Proteins

• As researchers have learned more about enzymes and proteins their hypothesis is better stated as the one gene –one polypeptide hypothesis

• We now know that genes provide the instructions for making proteins

– each gene is usually hundreds or thousands of nucleotides long

Making Proteins From DNA1. Transcription – the process by which

genetic information is copied from DNA to RNA

– results in a single stranded RNA molecule that is carried from the nucleus to the cytoplasm

The Stages of Transcription

PromoterTranscription unit

RNA polymerase

Start point

53

35

DNA

Making Proteins From DNA

Steps of Transcription

1. RNA polymerase binds to a specific region on the DNA called a promoter

The Stages of Transcription

PromoterTranscription unit

RNA polymerase

Start point

53

35

35

53

UnwoundDNA

RNAtranscript

Template strand of DNA

1

Initiation. After RNA polymerase binds to the promoter, the DNA strands unwind, and the polymerase initiates RNA synthesis at the start point on the template strand.

DNA

Making Proteins From DNA

Steps of Transcription

1. RNA polymerase binds to a specific region on the DNA called a promoter

2. This causes the DNA to separate

The Stages of TranscriptionPromoter

Transcription unit

RNA polymerase

Start point

53

35

35

53

53

35

5

Rewound

RNA

RNA

transcript

3

Unwound

DNA

RNA

transcript

Template strand of DNA

1 Initiation. After RNA polymerase binds to

the promoter, the DNA strands unwind, and

the polymerase initiates RNA synthesis at the

start point on the template strand.

2Elongation. The polymerase moves downstream, unwinding the

DNA and elongating the RNA transcript 5 3 . In the wake of

transcription, the DNA strands re-form a double helix.

DNA

Making Proteins From DNA

Steps of Transcription

1. RNA polymerase binds to a specific region on the DNA called a promoter

2. This causes the DNA to separate

3. RNA polymerase adds nucleotides one by one forming a new RNA molecule until it reaches a termination (or “stop”) signal

4. The newly formed strand is called mRNA

• mRNA – a single uncoiled chain of RNA nucleotides that carries genetic information to the ribosome for protein production

5. mRNA travels from the nucleus to the ribosome for protein synthesis

Making Proteins From DNA• Transcription Video

• Real Time Transcription Video

Making Proteins From DNA2. Translation – the process of making polypeptides

(proteins) using mRNA as a template

• the mRNA is converted into a sequence of amino acids, which makeup proteins

Making Proteins From DNA2. Translation – the process of making

polypeptides (proteins) using mRNA as a template

• the mRNA is converted into a sequence of amino acids, which makeup proteins

• the mRNA is broken up into segments of 3’s called codons

– codon – a sequence of 3 nitrogen bases in mRNA that code for an amino acid

Making Proteins From DNA• codon – a sequence of 3 nitrogen bases in mRNA

that code for an amino acid

– the sequence of codons along the mRNA is decoded into a sequence of amino acids which make up the polypeptide chain

Figure 17.4 The triplet code

DNAmolecule

Gene 1

Gene 2

Gene 3

DNA strand(template)

TRANSCRIPTION

mRNA

Protein

TRANSLATION

Amino acid

A C C A A A C C G A G T

U G G U U U G G C U C A

Trp Phe Gly Ser

Codon

3 5

35

Making Proteins From DNA

• Example:

• mRNA = CUGCUAGCUAGCUUCGAUCGAUGA

• a.a. = Leu- Leu- Ala- Ser- Phe- Asp - Arg- Stop

Summary

• Summary: DNA → RNA → Protein

• DNA = TACTCGATGGATTCGAACTCGATC

• RNA = AUGAGCUACCUAAGCUUGAGCUAG

• a.a = Meth- Ser- Tyr- Leu- Ser- Leu- Ser- stop

Transcription in More Detail

Transcription has 3 main steps:

1. Initiation

2. Elongation

3. Termination

Initiation

PromoterTranscription unit

RNA polymerase

Start point

53

35

Transcription in More Detail1. Initiation

– Each DNA sequence has a region called the promoter

• promoter – A region of DNA where RNA polymerase attaches and initiates transcription

– sometimes called a promoter gene

– After the promoter is recognized the RNA polymerase attaches to it

• RNA polymerase – an enzyme that adds nucleotides to the growing chain of RNA during transcription

– Once the polymerase is attached transcription can continue

Transcription in More Detail

2. Elongation

• RNA polymerase moves along the DNA and adds nucleotides to the growing RNA molecule

Transcription in More Detail

3. Termination

• Transcription continues until the RNA polymerase reaches a DNA sequence called a terminator

Preparing the RNA Sequence for Protein Production

• The RNA sequence made by transcription is not ready for translation. It must first be modified.

• Pre-mRNA modification, called Gene Splicing The average length of a RNA molecule made by transcription is 8000 nucleotides. However, it takes only 1200 nucleotides to code for the average protein.

• What does this tell you? It means that the mRNA has long nucleotide sequences that don’t code for genes

Preparing the RNA Sequence for Protein Production

• introns – a noncoding segment of nucleic acid that lie between coding segments of nucleic acids

• exons – coding segments of nucleic acids that are eventually translated into amino acids

Preparing the RNA Sequence for Protein Production

• The introns need to be cut from the RNA molecule and the exons joined together to form a mRNA molecule with a continuous coding sequence that can go on to translation

• spliceosome – molecule that cuts the introns out and joins exons back together

Translation

• We learned that DNA is converted into a long strand of pre-mRNA. The pre-mRNA strand is the then modified by gene splicing (cutting introns out) to get the mRNA strand that is carried to the cytoplasm for protein synthesis

• translation – the process of making polypeptides (proteins) using mRNA as a template

Translation

• Key terms to know

– codon – a sequence of 3 nitrogen bases in mRNA that code for a specific amino acid

• AUG – the start codon

– codon found on every functioning mRNA

– codes for the amino acid methionine

• UAA, UAG, or UGA – stop codons

Translation• Key terms to know

– rRNA – consists of RNA nucleotides that join proteins to make up the ribosome

Translation

• Key terms to know

– tRNA – a single strand of about 80 RNA nucleotides that carries amino acids to the ribosome for protein synthesis

Figure 17.14 The structure of transfer RNA (tRNA)3ACCACGCUUAA

GACACCU

GC

*G U G U

CUGAG

GU

A

AA G

UC

AGACC

C G A GA G G

G

GACUCA

UUUAGGCG5

Amino acidattachment site

Hydrogenbonds

AnticodonTwo-dimensional structure. The four base-paired regions and three loops are characteristic of all tRNAs, as is the base sequence of the amino acid attachment site at the 3 end. The anticodon triplet is unique to each tRNA type. (The asterisks mark bases that have been chemically modified, a characteristic of tRNA.)

(a)

*

*

**

*

**

*

***

Translation

• Key terms to know

– anticodon – a region on the tRNA consisting of 3 bases complementary to the codon of mRNA

tRNA

Amino acidattachment site

Hydrogen bonds

AnticodonAnticodon

(b) Three-dimensional structure

A A G

53

3 5

Symbol used in this book

(c)

Steps of Translation

1. Initiation

Steps of Translation

1. Initiation

• the ribosome binds to the mRNA at the start codon AUG

• the tRNA carrying the amino acid methionine binds to the mRNA strand where the start codon AUG is located

Figure 17.17 The initiation of translation

The arrival of a large ribosomal subunit completes the initiation complex. Proteins called initiationfactors (not shown) are required to bring all the translation components together. GTP provides the energy for the assembly. The initiator tRNA is in the P site; the A site is available to the tRNA bearing the next amino acid.

2

Initiator tRNA

mRNA

mRNA binding site Smallribosomalsubunit

Largeribosomalsubunit

Translation initiation complex

P site

GDPGTP

Start codon

A small ribosomal subunit binds to a molecule of mRNA. In a prokaryotic cell, the mRNA binding site on this subunit recognizes a specific nucleotide sequence on the mRNA just upstream of the start codon. An initiator tRNA, with the anticodon UAC, base-pairs with the start codon, AUG. This tRNA carries the amino acid methionine (Met).

1

MetMet

U A C

A U G

E A

3

5

5

3

35 35

Steps of Translation

•Elongation

Steps of Translation

2. Elongation

• amino acids are add one by one to the first amino acid methionine

• translocation – the process of mRNA moving along the ribosome making the polypeptide chain

– elongation continues to occur until the polypeptide is complete

Figure 17.18 The elongation cycle of translation

Amino endof polypeptide

mRNA

Codon recognition. The anticodon of an incoming aminoacyl tRNA base-pairs with the complementary mRNA codon in the A site. Hydrolysisof GTP increases the accuracy andefficiency of this step.

Ribosome ready fornext aminoacyl tRNA

Translocation. The ribosome translocates the tRNA in the A site to the P site. The empty tRNA in the P site is moved to the E site, where it is released. The mRNA moves along with its bound tRNAs,bringing the next codon to be translated into the A site.

Peptide bond formation. An rRNA molecule of the large 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 attaches the polypeptide to the tRNA in the A site.

E

P A

E

P A

E

P A

E

P A

GDPGTP

GTP

GDP

2

2

site site5

3

3

2

1TRANSCRIPTION

TRANSLATION

DNA

mRNARibosome

Polypeptide

Figure 17.19 The termination of translation

Release factor

Freepolypeptide

Stop codon(UAG, UAA, or UGA)

When a ribosome reaches a stop codon on mRNA, the A site of the ribosome accepts a protein called a release factor instead of tRNA.

The release factor hydrolyzes the bond between the tRNA in the P site and the last amino acid of the polypeptide chain. The polypeptide is thus freed from the ribosome.

1 2 3

5

3

5

3

5

3

The two ribosomal subunits and the other components of the assembly dissociate.

Steps of Translation

3. Termination

• elongation continues until a stop codon is reached

– UAA, UAG, or UGA don’t code for an amino acid

• the polypeptide is released from the ribosome

• the mRNA and ribosome disassemble

Figure 17.19 The termination of translation

Release factor

Freepolypeptide

Stop codon(UAG, UAA, or UGA)

When a ribosome reaches a stop codon on mRNA, the A site of the ribosome accepts a protein called a release factor instead of tRNA.

The release factor hydrolyzes the bond between the tRNA in the P site and the last amino acid of the polypeptide chain. The polypeptide is thus freed from the ribosome.

1 2 3

5

3

5

3

5

3

The two ribosomal subunits and the other components of the assembly dissociate.

Figure 17.13 Translation: the basic concept

TRANSCRIPTION

TRANSLATION

DNA

mRNARibosome

Polypeptide

Polypeptide

Aminoacids

tRNA withamino acidattached

Ribosome

tRNA

Anticodon

mRNA

Trp

Phe Gly

AG C

A A A

CC

G

U G G U U U G G C

Codons5 3

Translation Video

Real Time Translation

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

AUGCAGUCGAGGAUCAUGCGAUGAC

Intron Intron

pre-mRNA

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

AUGCAGUCGAGGAUCAUGCGAUGAC

Intron Intron

pre-mRNA

mRNA = A U G C A C G A G A U G C G A U G A C

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

AUGCAGUCGAGGAUCAUGCGAUGAC

Intron Intron

pre-mRNA

mRNA = A U G C A C G A G A U G C G A U G A C U A C

a.a. = Met

tRNA

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

AUGCAGUCGAGGAUCAUGCGAUGAC

Intron Intron

pre-mRNA

mRNA = A U G C A C G A G A U G C G A U G A C U A C G U G

a.a. = Met -- His

tRNA

Review

DNA = TACGTCAGCTCCTAGTACGCTACTG

AUGCAGUCGAGGAUCAUGCGAUGAC

Intron Intron

pre-mRNA

mRNA = A U G C A C G A G A U G C G A U G A C U A C G U G C U C U A C G C U A C U G

a.a. = Met -- His -- Glu -- Met -- Arg -- Stop

tRNA

Figure 17.23 The molecular basis of sickle-cell disease: a point mutation

In the DNA, themutant templatestrand has an A where the wild-type template has a T.

The mutant mRNA has a U instead of an A in one codon.

The mutant (sickle-cell) hemoglobin has a valine (Val) instead of a glutamic acid (Glu).

Mutant hemoglobin DNAWild-type hemoglobin DNA

mRNA mRNA

Normal hemoglobin Sickle-cell hemoglobin

Glu Val

C T T C A T

G A A G U A

3 5 3 5

5 35 3

Figure 17.24 Base-pair substitutionWild type

A U G A A G U U U G G C U A AmRNA5

Protein Met Lys Phe Gly Stop

Carboxyl endAmino end

3

A U G A A G U U U G G U U A A

Met Lys Phe Gly

Base-pair substitution

No effect on amino acid sequenceU instead of C

Stop

A U G A A G U U U A G U U A A

Met Lys Phe Ser Stop

A U G U A G U U U G G C U A A

Met Stop

Missense A instead of G

NonsenseU instead of A

Figure 17.25 Base-pair insertion or deletion

mRNA

Protein

Wild type

A U G A A G U U U G G C U A A5

Met Lys Phe Gly

Amino end Carboxyl end

Stop

Base-pair insertion or deletion

Frameshift causing immediate nonsense

A U G U A A G U U U G G C U A

A U G A A G U U G G C U A A

A U G U U U G G C U A A

Met Stop

U

Met Lys Leu Ala

Met Phe GlyStop

MissingA A G

Missing

Extra U

Frameshift causing extensive missense

Insertion or deletion of 3 nucleotides:no frameshift but extra or missing amino acid

3