25.1 dna structure and replication alfred hershey and martha chase (1952) –demonstrated that dna...

25.1 DNA Structure and Replication

• Alfred Hershey and Martha Chase (1952)– Demonstrated that DNA (deoxyribonucleic acid) is the

genetic material• Not proteins

– Two experiments• Viral DNA labeled with 32P found in the bacteria and not the

medium – it had entered the bacteria• Viral protein in capsids labeled with 35S found in medium and

not in the bacterium – it never entered the bacteria• Only the DNA entered the bacteria

Copyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

centrifuge

virus

1. When bacteria and viruses are cultured together, radioacti veviral DNA enters bacteria.

2. Agitation in blender dislodges viruses. Radioact vity stays inside bacteria.

Viruses inliquid are notradioactive.Bacteria insediment areradioactive.

3. Centrifugation separates viruses from bacteria and allows investigator to detect location of radioactivity.

capsid

centrifuge

DNAlabeledWith 32P

bacterium

capsidlabeledWith 32S

1. When bacteria and viruses are cultured together, radioactive viral capsids stay outside bacteria.

b. Viral capsid is labeled (yellow).

2. Agitation in blender dislodges viruses. Radioactivity stays outside bacteria.

3. Centrifugation separates viruses from bacteria and allows investigator to detect location of rdioactivity.

V iruses inliquid areradioactive.Bacteria insedimentare notradioactive.

a. Viral DNA is labeled (yellow).

Hershey-Chase Experiments


• Structure of DNA– James Watson and Francis Crick determined

the structure of DNA in 1953– DNA is a chain of nucleotides– Each nucleotide is a complex of three

subunits• Phosphoric acid (phosphate)• A pentose sugar (deoxyribose)• A nitrogen-containing base


• Structure of DNA– Four Possible Bases

• Adenine (A) - a purine• Guanine (G) - a purine• Thymine (T) - a pyrimidine• Cytosine (C) - a pyrimidine

– Complimentary Base Pairing• Adenine (A) always pairs with Thymine (T)• Guanine (G) always pairs with Cytosine (C)


P

SS

S

S

P

S

P

S

P

P

S

P

S

P

S

S

S

S

a. Double helix c. One pair of bases

P

S

P

P

P

P

P

OC

CC

purine basepyrimidine basephosphate3' end5' end

3' end5' end

5'1'

3' 2'

5'

4' 1'

1'

3' 2'

4'

2' 3'

5'

4'

deoxyribose

b. Ladder structure

3' end 5' end

C C

Overview of DNA Structure


• Replication of DNA– Semi-conservative replication

• Each daughter DNA molecule consists of one new chain of nucleotides and one from the parent DNA molecule

– The two daughter DNA molecules will be identical to the parent molecule


• Replication of DNA1. Before replication begins, the two strands of the parent

molecule are hydrogen-bonded together

2. Enzyme DNA helicase unwinds and “unzips” the double-stranded DNA

3. New complementary DNA nucleotides fit into place along separated strands by complementary base pairing. These are positioned and joined by DNA polymerase.

4. DNA ligase seals any breaks in the sugar-phosphate backbone

5. The two double helix molecules identical to each other and to the original DNA molecule


daughter DNA double helix

daughter DNA double helix

A

A

A

A

A

AA

A

AA

A

A

A

A

A

A

T

T

T

TT

T

T

T

TT

T

G

G

G

G

G

G

G

G

GG

G

G

C

CC

C

C

C

C

CC

C

5' 3'

G

A

5'

3'

region of parentalDNA double helix

region ofreplication:New nucleotidesare pairingwith those ofparental strands

region ofcompletedreplication

oldstrand

newstrand

newstrand

oldstrand

Overview of DNA Replication


Parental DNA molecule containsso-called old strandshydrogen-bonded by complementarybase pairing.

Region of replication. Parental DNAis unwound and unzipped. Newnucleotides are pairing with thosein old strands.

Replication is complete. Eachdouble helix is composed of an old(parental) strand and a new(daughter) strand.

Ladder Configuration and DNA Replication

25.2 RNA Structure and Function

• RNA (ribonucleic acid)– Uracil (U) used instead of thymine (T)

– Helper to DNA

– Three major types

G

U

A

C

S

S

S

S

P

P

P

P

base isuracil insteadof thymine

one nucleotideribose

Structure of RNACopyright © The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

Comparison of DNA and RNA

25.2 RNA Structure and Function

• Three Classes of RNA– Messenger RNA (mRNA)

• Produced in nucleus during transcription• Takes a message from DNA to the ribosomes

– Transfer RNA (tRNA)• Produced in nucleus• Transfers amino acids to ribosomes• Each tRNA carries only one type of amino acid

– Ribosomal RNA (rRNA)• Produced in nucleus• Makes up ribosomes (along with proteins)

25.3 Gene Expression

• Gene Expression Requires Two Steps1. Transcription

• Takes place in the nucleus• Portion of DNA serves as a template for mRNA formation

2. Translation• Takes place in the cytoplasm• Sequence of mRNA bases (which are complementary to

those in the template DNA) determines the sequence of amino acids in a polypeptide


• Transcription– During transcription, a gene (segment of the DNA)

serves as a template for the production of an RNA molecule

• All three classes of RNA

– Messenger RNA (mRNA)• RNA polymerase binds to a promoter (in DNA)• DNA helix is opened so complementary base pairing can

occur• RNA polymerase joins new RNA nucleotides in a sequence

complementary to that on the DNA


RNApolymerase

DNATemplatestrand

mRNAtranscript

To RNA processing

3'

5'

Transcription of DNA to Form mRNA


• Transcription– Processing of mRNA

• Primary mRNA contains bases complementary to both intron and exon segments of DNA

– Introns are intragene segments - removed– Exons are the portion of a gene that is expressed

• Intron sequences are removed• Guanine cap and poly-A tail added• Mature mRNA transcript ready


e ee i i

e ee i i

DNA

e ee

(cut-out) (cut-out)

DNA to be transcribed

transcription

poly-Atail

e = exonsi = introns

maturemRNA

primaryRNA

cap

mRNA Processing


• Translation– Gene expression leads to protein synthesis– The Genetic Code

• Triplet code - each three-nucleotide unit of a mRNA molecule is called a codon

• There are 64 different mRNA codons– 61 code for particular amino acids

• Redundant code - some amino acids have numerous code words

• Provides some protection against mutations– three are stop codons that signal polypeptide termination


Second Base

U C A G

U

C

A

G

U

C

A

G

U

C

A

G

U

C

A

G

U

C

A

G

ThirdBase

FirstBase

UUUphenylalanine

UUCphenylalanine

UUAleucine

UUGleucine

CUUleucine

CUCleucine

CUAleucine

CUGleucine

AUUisoleucine

AUCisoleucine

AUAisoleucine

AUG (start)methionine

GUUvaline

GUCvaline

GUAvaline

GUGvaline

GCGalanine

GCAalanine

GCCalanine

GCUalanine

ACGthreonine

ACAthreonine

ACCthreonine

ACUthreonine

CCGproline

CCAproline

CCCproline

CCUproline

UCGserine

UCAserine

UCCserine

UCUserine

UAUtyrosine

UGUcysteine

UGCcysteine

UACtyrosine

UGAstop

UAAstop

UGGtryptophan

UAGstop

CGUarginine

CAUhistidine

CGCarginine

CAChistidine

CGAarginine

CAAglutamine

CGGarginine

CAGglutamine

AGUserine

AAUasparagine

AGCserine

AGAarginine

AGGarginine

GGUglycine

GGCglycine

GGAglycine

GGGglycine

GAGglutamate

GAAglutamate

GACaspartate

GAUaspartate

AAGlysine

AAAlysine

AACasparagine

Messenger RNA Codons

• Transfer RNA– tRNA transports amino acids to the ribosomes– Boot-like shape– Amino acid binds to one end, the opposite end has an

anticodon• Triplet of three bases complementary to a specific

codon of mRNA– Order of mRNA codons determines the order in which

tRNA brings in amino acids



arginine

G

C

U

anticodon

anticodon

amino acid

a.

amino acid

b.

Transfer RNA: Amino Acid Carrier


G

G

CC

G

G

C

C

G

G

C G

C

G

C

G

C

C

G

A

U

T

A

A

T

T

A

U

T

G

C

G

DNA

mRNA

codon 1 codon 2 codon 3

anticodon 1GCC

anticodon 2 anticodon 3

polypeptide N N NC C C C C C

arginine

O O O

tryptophantyrosine

trnslationat ribosome

transcriptioninnucleus

DNAdouble helix

ACCAUG

R1 R2 R3

Overview of Gene Expression


• Ribosome and Ribosomal RNA– Free or attached to endoplasmic reticulum– Two subunits – one large and one small– Binding site for mRNA and three tRNAs– Binding sites facilitate complementary base pairing

between tRNA anticodons and mRNA codons– Brings amino acids in line in a specific order to form a

polypeptide– Several ribosomes may move along the same mRNA

• Multiple copies of a polypeptide may be made• The entire complex is called a polyribosome


mRNA

polypeptide

mRNA

large subunit

small subunit

a. Structure of aribosome

tRNA bindingsites

out goingtRNA

b . Binding sites of ribosome

incomingtRNA

c. Function of ribosomes d. Polyribosome

5' 3'

D: Courtesy Alexander Rich;

Polyribosome Structure and Function


• Translation Requires Three Steps– Initiation (requires energy)

• Initiation factors assemble components• Start codon (AUG)• P (peptide) site, A (amino acid) site, and E (exit)

site– Elongation (requires energy)– Termination

• Stop codon• Requires release factor


UC

A

U G G

3'5'

met

asp

ala

ser

3'5'

met

asp

ala

ser thr

3'5'

anticodon

tRNAmet

asp

ala

ser

3'5'

met

ala

ser

AUG

C U G

G A C AUG G A C

4. The ribosome moves for ward; the “empty” tRNA exits from the E site; the next amino acid–tRNA complex is approaching the ribosome.

3. Peptide bond formation attaches the peptide chain to the newly arrived amino acid.

2. Two tRNAs can be ta a ribosome at one time; the anticodons are paired to the codons.

Elongation

1. A tRNA–amino acid approaches the ribosome and binds at the A site.

peptidebond

tryp

val val

tryp

tryp

val

tryp

val

AUG G A C G A CAUG A C CC U GUC A C U GUC A UC A C U G

asp

Elongation


stop codon

UU

AG A

U G A

UC

UThe ribosome comes to a stopcodon on the mRNA. A releasefactor binds to the site.

release factor

The release factor hydrolyzes the bondbetween the last tRNA at the P site andthe polypeptide, releasing them. Theribosomal subunits dissociate.

Termination

3'

3'

5'

5'

asp

ala

tryp

val

glu

glu

ala

asp

tryp

val

AA U G A

Termination


DNA

mRNA

anticodon

tRNA

peptide

codonribosome

5'

C CG G

GCG

CG

C

CCC

GUA

CUA

UA

UA

UUA A

TRANSCRIPTION

3. mRNA moves into cytoplasm and becomes associated with ribosomes.

4. tRNAs with anticodons carry amino acids to mRNA.

5. Anticodon–codon complementary base pairing occurs.

6. Polypeptide synthesis takes place one amino acid at a time.

1. DNA in nucleus serves as a template for mRNA.

2. mRNA is processed before leaving the nucleus.

aminoacids

primarymRNA

int rons

nuclearpoer

TRANSLATION

3'5'

3'maturemRNA

large and smallribosomal subunits

Review of Gene Expression


Housekeeping

Hemoglobin

Insulin

Myosin

Gene type

Cell type Red blood Muscle Pancreatic

25.4 Control of Gene Expression• Housekeeping genes govern functions that are common

to many types of cells– Active in many cell types

• Only certain genes are active in cells that perform specialized functions

• Gene expression is controlled in a cell and this control accounts for its specialization

25.4 Control of Gene Expression

• Gene Expression in Prokaryotes– Operon: Cluster of genes usually coding for proteins

related to a particular metabolic pathway, along with the short DNA sequences that coordinately control their transcription

• Sequences consists of:– Promoter: A sequence of DNA where

transcription begins– Operator: A sequence of DNA where a

repressor protein binds


• Gene Expression in Bacteria

– Example: The lac Operon in E. coli

• When lactose is absent:– The regulator gene codes for a repressor that is normally

active

– A repressor protein binds to the operator

– RNA polymerase cannot transcribe the three structural

genes of the operon (the structural genes are not

expressed)


• Gene Expression in Bacteria– Example: The lac Operon in E. coli

• When lactose is present:– Lactose binds with the lac repressor– Repressor is unable to bind to the operator– Structural genes are transcribed

• Enzymes are produced

• lac operon is considered an inducible operon– Only activated when lactose induces its expression

• Repressible operons are usually active until a repressor turns them off


DNA

mRNA

lactose

DNA

RNA polymerase cannot bind to promoter.

regulator gene promoter operator structural genes

Active repressor

3'

act ive repressor

a. Lactose absent. Enzymes needed to metaboliz leactose are not produced.

5'

5'

3'5'

3' RNA polymerase can bind to promoter.

active repressor

b . Lactose present. Enzymes needed to met a bolize lactose are produced only when lactose is present.

enzymes forlactose metabolism

inact ive repressor

The lac Operon


• In bacteria:– Single promoter serves several genes that make up

a transcription unit• In eukaryotes, each gene has its own promoter• Bacteria rely mostly on transcriptional control• Eukaryotes employ a variety of mechanisms to

regulate gene expression– Affect whether the gene is expressed, the speed with

which it is expressed, and how long it is expressed


• Gene Expression in Eukaryotes– Levels of Gene Control

1. Pretranscriptional control

2. Transcriptional control

3. Posttranscriptional control

4. Translational control

5. Posttranslational control


functional protein

polypeptide chain

Posttranscriptional control

Transcriptional control

nuclear pore

intron exon

histones

nuclear envelope

Pretranscriptionalcontrol

primarymRNA

maturemRNA

Translationalcontrol

Posttranslationalcontrol

plasmamembrane

5´

3´

3´

5´

Levels of Gene Control in Eukaryotes


1. Pretranscriptional Control– Use DNA methylation and chromatin packing to keep

genes turned off– Heterochromatin: inactive genes located within darkly

staining portions of chromatin • Barr body• Tortoiseshell cats

– Euchromatin: loosely packed areas of active genes• Still needs to be “unpacked” before it can be transcribed• Chromatin remodeling complex pushes aside nucleosomes


© Photodisc/Getty RF;

active X chromosome

inactive X

inactive X

active X chromosome

cell division Barr bodies

Coats oftortoiseshellcats havepatches oforange andblack.

allele fororange color

allele forblack color

Females have twoX chromosomes.

One X chromosome is inactivatedin each cell. Which one of the pairis inactivated is random.


histone

nucleosome

Chromatinremodelingcomplex

DNA

X-Inactivation


2. Transcriptional Control– Transcription factors bind to promoter to form

transcription initiation complex– Transcription activators speed transcription

dramatically• Bind to enhancer

3. Posttranscriptional Control– Primary RNA processed

• Addition of poly-A tail and a guanine cap• Removal of introns and splicing of exons

– Different patterns of splicing can occur


4. Translational Control– Differences in the poly-A tails and/or guanine caps

may determine how long a mRNA is available for translation

– Specific hormones may also effect longevity of mRNA

5. Posttranslational Control– Some proteins must be activated after synthesis– Many proteins function only for a short time before

they are degraded or destroyed by the cell

25.5 Gene Mutations

• Permanent change in the sequence of bases in DNA

• Effect can range from none to complete inactivity of the protein

• Germ-line mutations can be passed to subsequent generations

• Some mutations can lead to cancer

25.5 Gene Mutations

• Three causes of mutations– Error in replication

• Proofreading by DNA polymerase usually takes care of this

– Mutagens• Radiation and certain organic chemicals• DNA repair enzymes constantly monitor and repair any

irregularities

– Transposons – jumping genes• Specific DNA sequences that move within and between

chromosomes• Sometimes alters neighboring gene expression• Likely all organisms (including humans) have them


Normal gene

a.

b. c.

Mutated gene

transposon

cannotcode forpurple

pigment

codes forpurple

pigment

C: © Mondae Leigh Baker;

25.5 Gene Mutations

• Effect of Mutations on Protein Activity– Point mutations involve a change in a single

DNA nucleotide• Possible change in a specific amino acid• May have no effect• May produce an abnormal protein

– Sickle cell

• May produce an incomplete protein


a.

No mutationC G T

His Leu Thr Pro Glu Glu

His Leu Thr Pro Glu Glu

His Leu Thr Pro Glu

His Leu Thr Pro Stop

A

A

A

(normal protein)His His

(incomplete protein)

Glu Stop

Glu Val

(abnormal protein)

Val

Val

Val

Val

Val

A C T G G A G GT C C CC TTGGA

C G TA C T G G GT C C CC TTGGAA

C G TA C T G G GT CC CCTGGAAG

C G TA C T G G GT C C CTGGAAG

3' 5'

T

b. Normal red blood cell c. Sickled red blood cell

© Stan Flegler/Visuals Unlimited;

25.5 Gene Mutations

• Frameshift Mutations– One or more nucleotides are either inserted or

deleted from DNA– Result can be a completely new sequence of

codons and a nonfunctional protein

THE CAT ATE THE RAT

THE ATA TET HER AT

25.5 Gene Mutations

• Nonfunctional Proteins– A single nonfunctioning protein can have a dramatic

effect on the phenotype, because enzymes are often a part of metabolic pathways

– PKU (phenylketonuria)• Phenylalanine can’t be broken down and builds up

causing mental impairment– Androgen insensitivity

• Faulty receptor makes cells unable to respond• Genetic male looks like a normal female

25.5 Gene Mutations

• Mutations can cause cancer– In the United States, the three deadliest forms

of cancer are lung cancer, colon and rectal cancer, and breast cancer

– Development of cancer involves a series of accumulating mutations that can be different for each type of cancer

– Carcinogenesis begins with the loss of tumor suppressor gene activity and/or the gain of oncogene activity


receptorstimulating growth factor

cytoplasm

transcription factor

nucleus

oncogene

plasmamembrane

protein thatoverstimulatesthe cell cycle

signalingpathway

25.5 Gene Mutations

• Although cancers vary greatly, they usually follow a common multistep progression

• Most cancers begin as an abnormal cell growth that is benign, or not cancerous, and usually does not grow larger

• Growth may become malignant, meaning that it is cancerous and possesses the ability to spread


epithelial cells 1 mutationCell (dark pink) acquires a mutation for repeated cell division.

New mutations arise, and one cell (green) has the ability to start a tumor.

The tumor is at itsplace of origin.One cell (purple)mutates further.

2 mutations

3 mutations tumor

lymphaticvessel

bloodvessel


Cells have gained the ability to invade underlying tissues by producing a proteinase enzyme.

Cancer cells nowhave the ability toinvade lymphaticand blood vessels.

New metastatictumors are foundsome distancefrom the tumor.

invasivetumor

malignanttumor

distant tumor

lymphaticvessel


epithelial cells 1 mutationCell (dark pink) acquires a mutation for repeated cell division.

New mutations arise, and one cell (green) has the ability to start a tumor.

The tumor is at itsplace of origin.One cell (purple)mutates further.

Cells have gained the ability to invade underlying tissues by producing a proteinase enzyme.

Cancer cells nowhave the ability toinvade lymphaticand blood vessels.

New metastatictumors are foundsome distancefrom the tumor.

2 mutations

3 mutations tumor

lymphaticvessel

bloodvessel

invasivetumor

malignanttumor

distant tumor

lymphaticvessel

Development of Cancer

25.5 Gene Mutations

• Characteristics of Cancer Cells– Cancer cells are genetically unstable– Cancer cells do not correctly regulate the cell

cycle– Cancer cells escape the signals for cell death– Cancer cells can survive and proliferate

elsewhere in the body

25.1 dna structure and replication alfred hershey and martha chase (1952) –demonstrated that dna...

Documents

dna replication25

daughter dna molecules

parent dna moleculethe

dna deoxyribonucleic

original dna moleculecopyright

radioacti veviral dna

viruses inliquid

rna structure