DNA

Griffith’s Experiment

Fredrick Griffith1928 British scientistWanted to see why people

got sick from bacteria (pneumonia)

Used mice and a strain of disease causing bacteriaHarmless strain had rough

edges when it was grown on a petri dish

Disease –causing strain had rough edges when grown on a petri dish

S-strain of bacteria killed mouse (smooth)R-strain does NOT kill mouse (rough)Heat kills the bacteria

What Griffith’s Experiment Proved

Griffith’s hypothesis:When live, harmless bacteria and heat-killed

bacteria are mixed, the heat-killed bacteria passed on disease-causing information to the live cells of the harmless bacteria, causing harmless bacteria to “transform” into bad bacteria

Transforming FactorThis factor was probably a gene because he

noticed that the offspring inherited the disease as well

Avery’s Experiments: DNA is the Transforming Factor

American biologist Oswald Avery, 1944 Was transforming factor DNA or Protein? Took Griffith’s heat-killed bacteria and good bacteria and

made an extract (aka a juice) Treated this extract “protein-destroying enzymes” Injected mice with the treated extract

Did the bacteria still function and kill the mice?Yes, the bad bacteria still transformed the harmless

bacteria…therefore protein did NOT contain the “transforming factor”

Treated Griffith’s deadly bacteria and harmless bacteria with “DNA-destroying enzymes”

Did they still function?No, the bacteria was not transformed, therefore, DNA had to

be the transforming factor

Avery’s ConclusionDNA is the cell’s genetic materialScientists were still skepticalProtein is made of 20 a.a. and DNA is only 4

nucleotides…didn’t make senseThey thought….”DNA is too simple!”

Hershey and Chase’s Experiments

1952, American biologists Alfred Hershey and Martha Chase

Which is the hereditary material: Protein or DNA ???

Conducted Experiments using viruses Viruses

Package of nucleic acid wrapped in a protein coatNot made of cellsCan only reproduce by infecting living cell with

its genetic infoGenetic info of virus then tells the cell’s

organelles to make more virusesBacteriophage (phage)-virus that infects

bacteria…literally means “bacteria eater” Virus attaches to the surface of bacteria, injects its genetic

material into the bacteria, the viral genes cause many more viruses to be made inside the bacteria until the bacteria burst and hundreds of new viruses are released

Hershey and Chase Experiment 1

Treated virus with radioactive sulfur-35 isotope (sulfur is in protein but not DNA)

Sulfur would attach to protein of virus If rad. sulfur was found in bacteria, that means it was the

protein coat of virus that contained hereditary information

Experiment 2 Treated virus with radioactive phosphorus-32 isotope

(phosphorus is in DNA but not protein) Phosphorus would attach to DNA of virus If rad. phosphorus was found in bacteria, that means it

was the protein coat of virus that contained hereditary information

Used blender to mix up bacteria and viruses Experiment 1=radioactive material was found out side

the bacteria cells Experiment 2=radioactive material was found inside

the bacterial cells

Hershey and Chase’s Experiments

Hershey and ChaseConclusion

The Phage’s (virus’) DNA entered the bacteria during infection but the protein did not

DNA must carry the genetic information of the virus

DNA is the hereditary material

DNA StructureDeoxyribonucleic

AcidDNA is a polymer

made up of many monomers called nucleotides

Nucleotide contains:5-carbon sugar

called deoxyriboseRNA contains RIBOSE

sugar insteadPhosphate groupOne Nitrogenous

base (there are 4 types)

Sugar and phosphate make up backbone (sides of ladder)

Nitrogenous bases make up steps of ladderBases are always paired (Chargaff’s rule)

What are these Nitrogenous bases???

Make up the “steps” of the DNA ladder One Step= A Purine + A

Pyrimidine

PurinesDouble ring structureAdenineGuanine

PyrimidinesSingle-ring structureCytosineThymine (in DNA only)Uracil (in RNA only)

Adenine (Purine) binds with Thymine (pyrimidine)TWO hydrogen bonds

Cytosine (Purine) binds with Guanine (pyrimidine)THREE hydrogen bonds

Chargaff’s RuleErwin Chargaff, Am.

biochemistYears before Watson

and CrickPercentage of guanine

and cytosine in DNA sample were about equal…the same with adenine and thymine

Chargaff’s Rule: A=T and G=C

Rule for “base-pairing”

Rosalind Franklin and Maurice Wilkins

Rosalind Franklin, 1950s, Brit scientist who studied DNA

Used X-ray diffraction technique to learn about structure of DNA

She noticed an X-shaped pattern

This indicated that DNA had a helical structure

She did not get a chance to publish her research…and her idea was basically stolen

Watson and Crick23-year old American

Biologist James Watson went to work with English Physicist Francis Crick

Used information from Rosalind Franklin and Maurice Wilkins (1950’s)These two scientists used

Rosalind Franklin’s x-ray crystallography images to begin to piece together the DNA structure

Showed that there were 2 strands twisted around each other

Watson and Crick

DNA is a double helixCreated the first accurate model of DNA

using wireBases paired up specifically with each other

b/c of hydrogen bondingComplementary base-pairing

A purine with a pyrimidineA=TG=C

DNA StructureChromatin wrapped

around proteins called histones

Chromatin condenses to form chromosomes during replication

This is what they already knew from the work of many scientists, about the DNA molecule:

DNA is made up of subunits which scientists called nucleotides.

Each nucleotide is made up of a sugar, a phosphate and a base.

There are 4 different bases in a DNA molecule: adenine (a purine) cytosine (a pyrimidine) guanine (a purine) thymine (a pyrimidine)

The number of purine bases equals the number of pyrimidine bases

The number of adenine bases equals the number of thymine bases

The number of guanine bases equals the number of cytosine bases

The basic structure of the DNA molecule is helical, with the bases being stacked on top of each other

Central Dogma of Biology

DNAmRNAproteinDNA TRANSCRIBES to mRNA

Process is called transcriptionmRNA TRANSLATES to

proteinsProcess is called translationmRNA actually makes amino

acids, which come together to make proteins

Nucleic AcidsDNA

Double strandDeoxyribose

sugarA=TG=C

RNASingle StrandRibose sugarA=UG=CUracil is the

nitrogenous base used instead of THYMINE

DNA parent strand makes 2 daughter strands…one fast, smart daughter strand (leading) and one, slower, no-fast daughter strand (lagging)

Leading strand (runs 3’ to 5’)

Lagging strand (runs 5’ to 3’)

How does DNA replicate itself?

Template mechanismWhat is a template???

PowerPoint presentations….Like the negative of a photograph

DNA ReplicationProcess of copying the DNA molecule

What phase of the CELL CYCLE?S-phase….

2 strands of double helix separate (Unzips)Each strand acts as a negative for making

the new complementary strandNucleotides line up one by one following

base pairing rulesEnzymes (DNA Polymerase and DNA

Ligase) link nucleotides together to form 2 new DNA strands called the daughter strands

DNA Replication SimplifiedUnzip parent DNAAdd nucleotides to the 2 template

strands of DNADNA parent strand makes 2 daughter strands…one

fast, smart daughter strand (leading) and one, slower, no-fast daughter strand (lagging)

Leading strand (runs 3’ to 5’)Lagging strand (runs 5’ to 3’)

Attach fragments on lagging strand

Origins of ReplicationSpecific site on DNA where

replication beginsDNA Helicase: enzyme that

binds to origin site and unwinds DNA in both directions

Copying goes outward in both directions making replication “bubbles”

Parent strands open up as daughter strands grow on both sides

Eukaryotic DNA has many origins of replication on a single DNA strandMakes copying faster

Eventually bubbles merge making two new strandsEach new strand has a part from

the original and a part from the new

Semi-conservative replicationMeans that parent of the original DNA

double helix is always kept as part of the new DNA copies

DNA PolymerasesEnzymesMake covalent bonds between nucleotides of

the new strandsFast, accurate process

Error only one in a billion nucleotides

Brings over nucleotides to unzipped DNA strand and drops them off

More Players in DNA Replication

DNA polymerase can only read a strand that is running 3-prime to 5-prime…DNA polymerase works non-stop

adding nucleotides onto the strand that runs in the 3’ to 5’ direction

Therefore, Only one strand is made by a smooth, and continuous process…

The other strand is put together in bits and pieces…Each little section of nucleotides

is called an “Okazaki Fragment”These are then “glued” together

to make one, continuous strand in the end by another enzyme…

DNA Ligase

Important Enzymes

DNA Helicaseunzips

DNA PolymeraseAdds nucleotides

DNA LigaseAttaches/glues

Central Dogma of Biology

DNAmRNAproteinDNA TRANSCRIBES to mRNA

Process is called transcriptionmRNA TRANSLATES to

proteinsProcess is called translationmRNA actually makes amino

acids, which come together to make proteins

DNAmRNAamino acids/polypeptide chain (Proteins)

DNA codes for an RNA strand The every 3 bases on the RNA

strand code for a specific amino acidCODON: three sequential

bases that code for a specific a.a. (20 a.a. total)

Amino acid are strung together to make a protein (primary structure)

Change DNA will change RNA which will change amino acids, which change protein

Legend: Transcription of DNA to RNA to protein: This dogma forms the backbone of molecular biology and is represented by four major stages.

1. The DNA replicates its information in a process that involves many enzymes: replication.

2. The DNA codes for the production of messenger RNA (mRNA) during transcription.

3. In eukaryotic cells, the mRNA is processed (essentially by splicing) and migrates from the nucleus to the cytoplasm.

4. Messenger RNA carries coded information to ribosomes. The ribosomes "read" this information and use it for protein synthesis. This process is called translation.

Ala: Alanine  Cys: Cysteine  Asp: Aspartic acid  Glu: Glutamic acidPhe: Phenylalanine  Gly: Glycine His: Histidine  Ile: Isoleucine Lys: Lysine Leu: Leucine  Met: Methionine Asn: AsparaginePro: Proline Gln: Glutamine Arg: Arginine Ser: SerineThr: Threonine Val: Valine Trp: Tryptophane Tyr: Tyrosisne

One Gene, One PolypeptideGeorge Beadle and Edward TatumAm. Geneticists1940sOrange bread mold, Neurospora

crassaStudied mutant strains of this mold

that could not growEach of the strains lacked a

specific enzyme needed to produce some molecule the mold needed to grow

Each strain was defective in a single, specific gene

“One-gene, one-enzyme” hypothesisThe function of an individual gene

is to dictate the production of specific enzyme

Scientists later learned that genes dictate not just enzymes, but a single polypeptide, so the hypothesis became known as the “One Gene, One Polypeptide” hypothesis

DNAmRNAProtein

TranscriptionDifferent form of the same

messageDNA makes single stranded

RNA (U replaces T)RNA leaves nucleus

TranslationTranslate from nucleic acid

language to amino acid language

Uses codons, 3-base “word” that codes for specific a.a.“code” for an amino acid

Several codons make a “sentence” that translates to a polypeptide (protein)

The Genetic CodeAm. Biochemist Marshall

Nirenberg began to crack the genetic code in the 1960sBuilt RNA model with uracil, called

poly U, conducted experiments with it and figured out UUU coded for amino acid phenylalanine

Scientists used his procedures to figure out the other amino acids represented by codons

Stop codons: UAA, UGA, UAGSIGNAL END OF GENETIC MESSAGE

Start codon: AUGSIGNAL TO START TRANSLATING an

RNA transcript

Start Codons

UAAUGAUAG

Stop Codons

AUG

Three Types of RNAmRNAtRNArRNA

Three Types of RNA… #1mRNA (messanger RNA)

RNA transcribed from DNA templateRNA polymerase (enzyme) links RNA nucleotides

togetherModified in nucleus before if exits

RNA splicing: process in which Introns are removed and exons re joined together to make a continuous coding mRNA molecule

IntronsInternal non-coding regions of DNA and mRNASpace fillersThey are cut out of mRNA before it is allowed to leave the

nucleusProcess is called RNA splicing (processing)

ExonsCoding region of DNA and mRNA that will be translated

(Expressed)VERY important part of mRNA…it is carrying the message from

DNA (def can’t cut this out)

Three Types of RNA…#2tRNA (transfer RNA)

The interpreterTranslate 3-letter base

codes into amino acids Carries anti-codon on one

end (three letters opposite of what is on mRNA)

Carries amino acid on other end

Anti-codon recognizes codon and attaches

Three Types of RNA…#3rRNA (ribosomal RNA)

Found in ribosomeRibosome composed of 2

subunits:Small subunit for mRNA to

attachLarge Subunit for two tRNAs

to attach“P” site: holds the tRNA

carrying the growing polypeptide chain

“A” site: holds the tRNA that is carrying the next a.a. to be added to the chain

When stop codon (UAA, UAG, UGA) is reached, translation ends and polypeptide is released from tRNA by hydrolysis