CHAPTER 9CHAPTER 9

DNA, RNA, Translation and DNA, RNA, Translation and TranscriptionTranscription

The Discovery Of DNA

Mendel studied the pea plants in the late 1800’s

Scientists wanted to know what it was that contained hereditary factors

An epidemic of Pneumonia in London in the 1920’s sparked research which eventually leads to the discovery of DNA.

GRIFFITH’s EXPERIMENTSGRIFFITH’s EXPERIMENTS

1928, London, England: Fredrick 1928, London, England: Fredrick Griffith was attempting to discover a Griffith was attempting to discover a vaccine for the virulent strain, or vaccine for the virulent strain, or disease causing strain of the disease causing strain of the Streptococcus pneumoniae Streptococcus pneumoniae bacteriumbacterium..

Streptocuccus pneumoniae was Streptocuccus pneumoniae was causing pneumoniacausing pneumonia

GRIFFITH’s EXPERIMENTGRIFFITH’s EXPERIMENT

By studying the bacterium he discovered By studying the bacterium he discovered that:that:

The virulent form was surrounded by a The virulent form was surrounded by a capsule made of polysaccarides that capsule made of polysaccarides that protects it from the body’s defenses. So, protects it from the body’s defenses. So, when grown in a petri dish, they grow as when grown in a petri dish, they grow as smooth colonies. (S strain)smooth colonies. (S strain)

There was an another strain that did not There was an another strain that did not cause pnuemonia. This harmless strain cause pnuemonia. This harmless strain lacked a capsule and as a result grew into lacked a capsule and as a result grew into rough colonies in the petri dish. (R strain)rough colonies in the petri dish. (R strain)

GRIFFITH’s EXPERIMENTSGRIFFITH’s EXPERIMENTS

Griffith experimented with mice.Griffith experimented with mice. He injected a mouse with the S strain and He injected a mouse with the S strain and

it diedit died He injected another mouse with the R He injected another mouse with the R

strain, it livedstrain, it lived He heated-killed the S strain and injected He heated-killed the S strain and injected

it into the mouse, it livedit into the mouse, it lived He then combined heat-killed S strain and He then combined heat-killed S strain and

live R strain and injected it into the mouse.live R strain and injected it into the mouse.

GRIFFITH’s RESULTSGRIFFITH’s RESULTS

When injected with the heat killed S When injected with the heat killed S strain combined with the R strain, the strain combined with the R strain, the mouse died.mouse died.

Griffith concluded that transformation Griffith concluded that transformation occurred.occurred.

TransformationTransformation is a type of transfer is a type of transfer of genetic material from one cell to of genetic material from one cell to another OR from one organism to another OR from one organism to another organism.another organism.

Avery’s Experiments

1940’s, United States: Oswald Avery wanted to test whether the transforming agent in Griffith’s experiments was protien, RNA or DNA.

The scientists used enzymes to destroy separately each of the 3 molecules in the Heat-Killed S strain.

Avery’s Experiments

Experiment #1: Protease was used to destroy the protien in one batch of the heat-killed S strain.

Experiment #2: RNase was used to destroy the RNA in another batch of heat-killed S Strain

Experiment #3: DNase was used to destroy the DNA in another batch of heat-killed S strain.

Each batch was combined with a batch of live harmless R strain and injected into different mice

Avery’s Results

The strain missing the protein and the RNA were still able to transform the R strain into the S strain and kill the mice.

The strain missing DNA did NOT transform the R strain and the mouse lived.

Result: DNA was responsible for transformation in the bacteria

HERSHEY-CHASE EXPERIMENTHERSHEY-CHASE EXPERIMENT 1952, United States: Martha Chase & 1952, United States: Martha Chase &

Alfred Hershey tested whether DNA or Alfred Hershey tested whether DNA or protein was the hereditary material protein was the hereditary material viruses transfer when viruses enter a viruses transfer when viruses enter a bacterium.bacterium.

BacteriophagesBacteriophages – viruses that infect – viruses that infect bacteriabacteria

HERSHEY-CHASE EXPERIMENTHERSHEY-CHASE EXPERIMENT

Radioactive isotopes was used to label the Radioactive isotopes was used to label the protein and DNA in the bacteriophage. protein and DNA in the bacteriophage.

Radioactive Sulfur (Radioactive Sulfur (3535S) was used to label S) was used to label proteinprotein

Radioactive phosphate (Radioactive phosphate (3232P) was used to P) was used to label DNAlabel DNA

Then, they allowed the protein-labeled and Then, they allowed the protein-labeled and the DNA-labeled bacteriophages to the DNA-labeled bacteriophages to independently infect E-coli bacteria.independently infect E-coli bacteria.

HERSHEY-CHASE RESULTSHERSHEY-CHASE RESULTS The scientists removed the coats of the The scientists removed the coats of the

viruses by putting the solution in a blenderviruses by putting the solution in a blender The scientists then separated the The scientists then separated the

bacteriophages from the E coli by bacteriophages from the E coli by centrifuge centrifuge

They discovered that all of the viral DNA They discovered that all of the viral DNA and only a little bit of the protein had and only a little bit of the protein had entered the E coli. entered the E coli.

ConclusionConclusion: DNA is the hereditary : DNA is the hereditary molecule in Virusesmolecule in Viruses

SECTION 2: DNA STRUCTURESECTION 2: DNA STRUCTURE

1950’s: By now most scientists 1950’s: By now most scientists understood and accepted that DNA was understood and accepted that DNA was molecule that contained the hereditary molecule that contained the hereditary information.information.

What they wanted to discover was how What they wanted to discover was how this molecule could replicate, store & this molecule could replicate, store & transmit hereditary informationtransmit hereditary information

THE DOUBLE HELIXTHE DOUBLE HELIX

1950’s, James Watson, Maurice Wilkens 1950’s, James Watson, Maurice Wilkens and Francis Crickand Francis Crick

Won the Nobel Prize in Medicine for their Won the Nobel Prize in Medicine for their theory about the structure of DNA.theory about the structure of DNA.

Structure was developed with the help of Structure was developed with the help of Rosalind Franklin and her x-ray Rosalind Franklin and her x-ray photographs of DNA crystalsphotographs of DNA crystals

DNA NUCLEOTIDESDNA NUCLEOTIDES

DNADNA is made up of is made up of TWOTWO long chains (or long chains (or strands) of repeating subunits called strands) of repeating subunits called nucleotidesnucleotides

NucleotidesNucleotides contain contain THREETHREE parts: parts: – DeoxyriboseDeoxyribose: A 5-carbon sugar: A 5-carbon sugar– A phosphate groupA phosphate group– A nitrogen baseA nitrogen base – contains a Nitrogen, – contains a Nitrogen,

Carbon and accepts hydrogen: Adenine, Carbon and accepts hydrogen: Adenine, Guanine, Cytosine, and ThymineGuanine, Cytosine, and Thymine

Bonds Hold DNA togetherBonds Hold DNA together The DNA double helix is similar to a The DNA double helix is similar to a

spiral ladder.spiral ladder. The alternating sugar and phosphate The alternating sugar and phosphate

molecules form the sides of the ladder molecules form the sides of the ladder Nucleotides are held together by Nucleotides are held together by

covalent bonds between the sugar of covalent bonds between the sugar of one nucleotide and the phosphate of one nucleotide and the phosphate of the next nucleotidethe next nucleotide

There are 10 nucleotide pairs for each There are 10 nucleotide pairs for each spiral turn of the DNA helixspiral turn of the DNA helix

Bonds Hold DNA together: the rungsBonds Hold DNA together: the rungs The bases (nitrogen bases) face toward The bases (nitrogen bases) face toward

the center of the DNA molecule.the center of the DNA molecule. The bases form hydrogen bonds with The bases form hydrogen bonds with

bases on the other side and make up bases on the other side and make up “the rungs of the ladder”“the rungs of the ladder”

All pairs are of uniform width: in each All pairs are of uniform width: in each pair, one base has a two ring structure pair, one base has a two ring structure and other base has a one ring structure and other base has a one ring structure

Nitrogen BasesNitrogen Bases

While the sugar and phosphate groups While the sugar and phosphate groups are identical, the nitrogen bases could are identical, the nitrogen bases could be one of four kind (broken down to be one of four kind (broken down to two groups by structure):two groups by structure):

Purines: contain a double ring of carbon Purines: contain a double ring of carbon & nitrogen: Adenine & Guanine& nitrogen: Adenine & Guanine

Pyrimidines: contain a single ring of Pyrimidines: contain a single ring of carbon & nitrogen: Thymine & Cytosinecarbon & nitrogen: Thymine & Cytosine

COMPLIMENTARY BASESCOMPLIMENTARY BASES Just before DNA structure was confirmed: it was Just before DNA structure was confirmed: it was

found that the percentage of adenine is equal to the found that the percentage of adenine is equal to the percentage of thyminepercentage of thymine

It was also found that the percentage of cytosine It was also found that the percentage of cytosine was equal to the percentage of guaninewas equal to the percentage of guanine

Upon Observation, Base-Pairing Rules was Upon Observation, Base-Pairing Rules was uncovered – cytosine pairs only with guanine, and uncovered – cytosine pairs only with guanine, and adenine pairs only with thymineadenine pairs only with thymine

COMPLIMENTARY BASESCOMPLIMENTARY BASES These pairs are known as Complementary These pairs are known as Complementary

base pairs – one double-ringed purine and base pairs – one double-ringed purine and one single-ringed pyrimidineone single-ringed pyrimidine

Because of these base pairing rules, the Because of these base pairing rules, the order if bases on one side of a chain of order if bases on one side of a chain of DNA is complementary to the order of DNA is complementary to the order of bases on the other side.bases on the other side.

Base Sequence: the order of nitrogen Base Sequence: the order of nitrogen bases on a chain of DNAbases on a chain of DNA

Complementary BasesComplementary Bases

WHY IS THE BASE PAIRING IMPORTANT?WHY IS THE BASE PAIRING IMPORTANT? Hydrogen bonds b/t the base pairs help hold the Hydrogen bonds b/t the base pairs help hold the

two strands of a DNA molecule togethertwo strands of a DNA molecule together The complementary nature of DNA helps explain The complementary nature of DNA helps explain

how it is that DNA replicates before a cell divides how it is that DNA replicates before a cell divides – One strand of a DNA molecule can serve as a – One strand of a DNA molecule can serve as a template for making a new complimentary template for making a new complimentary strand.strand.

DNA MODELSDNA MODELS You will see the structure of DNA simplified You will see the structure of DNA simplified

from the actual double helix model to a from the actual double helix model to a straight ladder model or just the base straight ladder model or just the base pairingspairings

This is because the only variable is the base This is because the only variable is the base pairs, the sugar and phosphate groups are pairs, the sugar and phosphate groups are identical in all DNAidentical in all DNA

Section 3: DNA REPLICATIONSection 3: DNA REPLICATION

The discovery of the double helix structure The discovery of the double helix structure of DNA explains how it can replicate exactly of DNA explains how it can replicate exactly each time a cell divides, the key feature of each time a cell divides, the key feature of hereditary material.hereditary material.

How DNA replicatesHow DNA replicates

DNA Replication is the process by which DNA Replication is the process by which DNA is copied in a cell before a cell divides DNA is copied in a cell before a cell divides by mitosis, meiosis or binary fission.by mitosis, meiosis or binary fission.

Because the two strands of DNA are Because the two strands of DNA are complimentary, each serve as a template to complimentary, each serve as a template to make a NEW COMPLIMENTARY STRANDmake a NEW COMPLIMENTARY STRAND

After replication, the 2 identical double-After replication, the 2 identical double-stranded DNA molecules separate & move stranded DNA molecules separate & move to new cells formed during cell division.to new cells formed during cell division.

STEPS OF DNA REPLICATIONSTEPS OF DNA REPLICATION

Step 1Step 1 Helicases: Enzymes that separate the DNA Helicases: Enzymes that separate the DNA

strandsstrands Helicase move along the strands and Helicase move along the strands and

breaks the hydrogen bonds between the breaks the hydrogen bonds between the complimentary nitrogen basescomplimentary nitrogen bases

Replication Fork: the Y shaped region that Replication Fork: the Y shaped region that results from the separation of the strandsresults from the separation of the strands

STEPS..STEPS.. Step 2Step 2 DNA Polymerase: enzymes DNA Polymerase: enzymes that ADD complimentary that ADD complimentary nucleotides.nucleotides. Nucleotides are found floating Nucleotides are found floating freely inside the nucleusfreely inside the nucleus Covalent bonds form between Covalent bonds form between the phosphate group of one the phosphate group of one nucleotide and the deoxyribose nucleotide and the deoxyribose of another of another Hydrogen bonds form between the complimentary nitrogen Hydrogen bonds form between the complimentary nitrogen

basesbases

STEPS..STEPS.. Step 3Step 3 DNA polymerases finish replicating the DNA DNA polymerases finish replicating the DNA

and fall off.and fall off. The result is two identical DNA molecules The result is two identical DNA molecules

that are ready to move to new cells in cell that are ready to move to new cells in cell division.division.

Semi-Conservative Replication: this type of Semi-Conservative Replication: this type of replication where one strand is from the replication where one strand is from the original molecule and the other strand is neworiginal molecule and the other strand is new

http://www.johnkyrk.com/DNAreplication.html

Something about Step 2Something about Step 2 In step 2, remember that each strand is In step 2, remember that each strand is

making its own new strand.making its own new strand. DNA synthesis is occurring in two different DNA synthesis is occurring in two different

directionsdirections One strand is being made towards the One strand is being made towards the

replication fork and the other is being made replication fork and the other is being made away from the fork. The strand being made away from the fork. The strand being made away from the fork has gaps.away from the fork has gaps.

Gaps are later joined by another enzyme, Gaps are later joined by another enzyme, DNA ligaseDNA ligase

http://www.lewport.wnyric.org/JWANAMAKER/animations/DNA%20Replication%20-%20long%20.html

Prokaryotic ReplicationProkaryotic Replication

Prokaryotic Cells have one circular Prokaryotic Cells have one circular chromosome. chromosome.

Two replication forks are formed in the same Two replication forks are formed in the same area of the chromosome.area of the chromosome.

They proceed in opposite directions – like They proceed in opposite directions – like two zipperstwo zippers

Replication continues along each fork until Replication continues along each fork until they meet and the entire the entire molecule they meet and the entire the entire molecule is copiedis copied

DNA ERRORSDNA ERRORS

Usually DNA replication occurs without any errors. Usually DNA replication occurs without any errors. Only about one error in about every billion Only about one error in about every billion replications occurs.replications occurs.

DNA polymerases have a repair function that DNA polymerases have a repair function that “proofreads” the DNA. It will replace a wrongly “proofreads” the DNA. It will replace a wrongly placed nitrogen baseplaced nitrogen base

MutationMutation: a change in the nucleotide sequence.: a change in the nucleotide sequence. A mutation can have serious effects on the A mutation can have serious effects on the

function of an important gene and disrupt an function of an important gene and disrupt an important cell functionimportant cell function

DNA ErrorsDNA Errors

Chemicals and Ultraviolet radiation from the sun Chemicals and Ultraviolet radiation from the sun and tanning booths can change DNAand tanning booths can change DNA

Some mutations can lead to cancer. Mutations in Some mutations can lead to cancer. Mutations in the way the cell divides can lead to tumors.the way the cell divides can lead to tumors.

An effective mechanism for the repair of damaged An effective mechanism for the repair of damaged DNA is very important to the survival of an DNA is very important to the survival of an organism.organism.

Studying DNA replication is important to Studying DNA replication is important to understanding and treating various types of understanding and treating various types of cancer.cancer.

Section 4: Protein SynthesisSection 4: Protein Synthesis

DNA contains genes that code for a DNA contains genes that code for a hereditary characteristic, example: hair hereditary characteristic, example: hair color.color.

The gene that codes for hair color directs The gene that codes for hair color directs the making of a protein called melanin in the making of a protein called melanin in the hair follicle. the hair follicle.

The protein is made through an The protein is made through an intermediate (middle man) – a nucleic acid intermediate (middle man) – a nucleic acid called RNA, Ribonucleic Acidcalled RNA, Ribonucleic Acid

RNA STRUCTURE & FUNCTIONRNA STRUCTURE & FUNCTION

DNA and RNA are similar in that they are DNA and RNA are similar in that they are both made up of nucleotides.both made up of nucleotides.

DNA and RNA differ in Four Ways:DNA and RNA differ in Four Ways:1.1. RNA has ribose, DNA has deoxyriboseRNA has ribose, DNA has deoxyribose2.2. RNA contains a nitrogen base uracil RNA contains a nitrogen base uracil

instead of thymineinstead of thymine3.3. RNA is single stranded*RNA is single stranded*4.4. RNA is much shorter than DNA. It RNA is much shorter than DNA. It

contains the information for one gene. contains the information for one gene.

TYPES OF RNATYPES OF RNA

Three Major Types of RNAThree Major Types of RNA

1.1. Messenger RNA (mRNA) ~ single Messenger RNA (mRNA) ~ single stranded RNA molecule that carries the stranded RNA molecule that carries the instructions from a gene to make a instructions from a gene to make a protein. Carries the genetic “message” protein. Carries the genetic “message” from the DNA in the nucleus to the from the DNA in the nucleus to the ribosomes in the cytosolribosomes in the cytosol

TYPES OF RNA

Three Major Types of RNA

2. Ribosomal RNA (rRNA) ~ part of the structure of ribosomes, where protein synthesis occurs

3. Transfer RNA (tRNA) ~ transfers amino acids to the ribosome to make a protein.

FLOW OF GENETIC INFORMATION

How the information goes from DNA to the Ribosomes and into protein form.

1. Transcription – DNA acts as a template for the synthesis of RNA

2. Translation – RNA directs the assembly of the proteins.

3. Protein Synthesis – proteins are formed based on information in DNA and carried out by RNA in the ribosomes

TRANSCRIPTION Transcription means that the information within

DNA is transcribed/“rewritten” as an RNA molecule

Occurs in three steps:1. RNA polymerase, an enzyme that catalyzes

(starts) the formation of RNA on a DNA template.

a. A promoter is a specific nucleotide sequence of DNA where RNA polymerase binds and initiates transcription.

b. After RNA polymerase binds to the promoter, DNA strands unwind and separate

TRANSCRIPTION

THREE STEPS OF TRANSCRIPTION

2. RNA polymerase adds free RNA nucleotides that are complementary to the nucleotides on one of the DNA strands. The resulting chain is an RNA molecule.

a. Complementary base-pairing determines the nucleotide sequence in the newly made RNA.

b. Transcription only occurs in a specific area (one gene) of the DNA. RNA polymerase moves past the area and DNA rewinds

TRANSCRIPTION

THREE STEPS OF TRANSCRIPTION3. RNA polymerase reaches the terminal

signal, a specific sequence of nucleotides that marks the end of the gene.

a. Upon reaching this mark, RNA polymerase releases both the DNA & the newly formed RNA.

b. The newly formed RNA can be any type of RNA, free to perform it’s job within the cell.

The Genetic Code

• What makes up a protein?– Amino Acids

• Instructions on which amino acids to assemble are coded within the sequence of nucleotides.

• Genetic Code – the term for the rules that relate how a sequence of nitrogen bases corresponds to a particular amino acid

• There are 20 different amino acids found in living things

The Genetic Code• Codon – each 3-nucleotide sequence in

mRNA that encodes an amino acid or signifies a start or stop sequence

• RNA Polymerase unwindsunwinds and unzipsunzips DNA(but does not proof-read… why not?)

• Complementary NTP’s add to templateDNA strand from 5’ to 3’

• RNA Polymerase begins transcribing the DNA at a specific point

• RNA strand is identical to the non- coded DNA (and complementary to the template strand)

EXCEPT FOR...

• Same processSame process as Prokaryotes!

• After mRNA is transcribed from DNA then the mRNA has a different fate in prokaryotes and eukaryotes

• Prokaryotes immediately begin translatingtranslating the mRNA. Eukaryotes must process it first.

mRNA Processing:• intron/exon• methyl cap• poly-A tail

No mRNA Processing

• Viral DNA injected into cells• Cells evolve nucleasesnucleases in cytoplasm that chomp up

any RNA or DNA out there• Nucleases can’t get through the nuclear envelope so

DNA is safe• mRNA sent out into the cytoplasm must be protected

– Methyl cap is a block– Poly A tail is a fuse

• mRNA is still chomped up into NTP’s and recycled, but the Poly A tail gives it some time

• Eukaryotic DNA is composed mostly of “non-coding “non-coding DNA”DNA” (or “junk DNA”)– We’re still not entirely sure what it does– Was probably inserted by different viruses over time– The ultimate selfish gene just hitching a ride on a successful

group of genes…

• The intronsintrons are the sections of DNA not expressed, the exonsexons are the sections that are expressed (ex-ons are ex-pressed, get it?)

• SpliceosomeSpliceosome loops out the introns and snips them out• So now we’ve got some mRNA that codes for a protein

TRANSLATION the making of a protein

The start codon: AUG – a specific sequence (Adenine, Uracil, Guanine) of nucleotides in mRNA that indicates where translation should begin – Methionine

UGA – stop codon - doesn’t code for an amino acid but signals for translation to stop

TRANSLATION

Every protein is made up of one or more polypeptides.

Polypeptides are amino acid chains which are linked by peptide bonds

Each polypeptide chain may consist of hundreds of thousands of the 20 different amino acids – arranged in a sequence that is specific to THAT PARTICULAR PROTEIN

TRANSLATION

STEP ONE Initiation: tRNA and mRNA join

together. The tRNA carries a anticodon – three nucleotides that are complimentary to the sequence of codon on the RNA (the start codon)

The start amino acid is methinonine but this would be removed later

TRANSLATIONSTEP TWO Elongation – the chain is put

together Another tRNA carrying the

second appropriate amino acid pairs with the mRNA

A peptide bond forms between the first amino acid and this one (and so on). The first tRNA detaches

TRANSLATION

Step Three The polypeptide chain continues to

grow Another tRNA moves in, carrying the

amino acid for the next mRNA codon The growing chain moves from one tRNA

to the amino acid attached to the next tRNA

TRANSLATION

Step Four The ribosome reaches the stop codon The newly made polypeptide falls off.Step Five The ribosome complex falls apart. The newly

made polypeptide chain is released The last tRNA leaves the ribosome The ribosome is free to translate the same

or another mRNA

THE HUMAN GENOME The Human Genome Genome – the complete genetic content In 1999, scientists have mapped the entire

gene sequence of the human genome They now know the order of the 3.2 billion

base pairs in the 23 human chromosomes The next challenge is to learn what

information the DNA sequences actually code for.