DNA

Chapter 12

Sex-linked trait

• Some traits can only be carried on an X chromosome.

• An example is red/green colorblindness.• Much more common in males, because males

only get on X chromosome.• A male with the recessive trait can only pass it on

to daughters.• Why?• The sons get the Y chromosome.

Frederick Griffith

• Originally was trying to investigate bacteria and how it causes pneumonia.

• He isolated two different strains of the bacteria.

• Only one strain caused pneumonia.• Injected mice with disease causing bacteria

and these mice died.• Injected mice with non-disease causing

bacteria and they did not die.

Griffith

• Injected mice with heat-killed, disease causing bacteria, and they did not die.

• Injected mice with heat-killed, disease causing bacteria and non-disease causing bacteria, and these mice died.

• Why?• He knew the heat-killed, disease causing

bacteria passed their disease causing ability to the non-disease causing bacteria.

Griffith

• He called this transformation.• One strain was apparently changed into

another.• Some factor had to be transferred from the

heat killed cells to the live cells.• Must contain a gene with info to change the

bacteria.

Avery Repeated Griffith’s work to isolate the molecule

that caused transformation. This could be what carried genes! They extracted a sample from the heat killed

bacteria. Inserted enzymes that destroyed lipids, proteins,

RNA, and carbs-transformation still occurred. Transformation did not occur when enzymes

broke down DNA.

Avery

• Avery and other scientists at that point could conclude:

• DNA is the nucleic acid that stores and transmits the genetic information from one generation of an organism to the next.

• This happened in 1944.

Hershey-Chase

• Two scientists studied viruses in 1952.• Bacteriophage-a virus that attacks and kills

bacteria.• Kills bacteria by injecting its DNA.• Consisted of a protein coat and a DNA core.• These bacteriophages multiply inside the bacteria,

eventually causing the cell to burst, releasing hundreds of bacteriophages.

• Look at page 289.

Bacteriophages

• Refer to page 290.• Injected bacteriophage DNA with radioactive

phosphorous.• Injected radioactive sulfur into protein coat.• Radioactivity showed up in the bacterium that

was infected with DNA radioactivity, not protein.

• Conclusion: genetic material in bacteriophage was from DNA.

What do genes do?

• Carry info from one generation to the next• Put that information to work by determining

the heritable characteristics of organisms• Had to be easily copied because of cell

division.• How does DNA do all of this?

Nucleotide

• Made of three parts:• 5 carbon sugar• Phosphate group• Nitrogenous base

Nitrogenous bases

• Purines: double-ringed• Adenine• Guanine

• Pyrimidines: single-ringed• Thymine • Cytosine

DNA

• Said to have a sugar-phosphate backbone.• The rungs of the ladder structure are the

bases.• Chargaff’s Rules:• A bonds with T=double hydrogen bonds• C bonds with G=triple hydrogen bonds• How do nucleotides code for anything?

Rosalind Franklin

• Used X-ray diffraction.• Shoots X-rays off of DNA to see how they

bounce back.• She revealed that DNA was double-stranded

and that it twisted in a helix shape.• She also determined that the bases were in

the middle.

Crick and Watson

• Tried to build 3-D models of DNA.• Used Franklin’s work combined with what

they had already figured out.• Published results in 1953.• Model was a double-helix that were wound

around each other.

Crick and Watson

• Why could Franklin not figure it out?• Watson and Crick found that hydrogen

bonds could barely hold bases together.• This provided the structural support for

DNA.• This principle is called base-pairing.

Prokaryotic DNA

• Do not have organelles or nuclei.• DNA exists in the cytoplasm-nowhere else to

go.• It is called the chromosome of the prokaryotic

cell.

Eukaryotic DNA

• Found in the nucleus.• When the cell is not dividing-in the form of

chromatin.• When the cell is dividing-in the form of

chromosomes.• Chromosomes are DNA plus proteins called

histones.

Size of DNA

• It must be very folded and compact.• From book:• Bacteria may contain 4.6 million base pairs.• The bacteria is only 1.6 µm. This means the

DNA must be folded very tightly.• Try to pack a rope that is 300 m long into a

backpack.

Eukaryotic DNA length

• Must be packed more tightly.• Contains 1000 more base pairs than

prokaryotes.• Each human cell contains more than one

meter of DNA.• Smallest human chromosome contains 30

million base pairs.

Chromosomes

• Tightly wound containing DNA and histones.• This forms a nucleosome.• Nucleosomes coil up and eventually become

supercoils, the tightest than can be wound.• Nucleosomes are very unique to be able to fold in

this way.• Histones have changed little over time.• DNA starts to be read by opening up

nucleosomes.

Replication

• In Watson and Crick’s paper, they theorized as to how DNA could replicate.

• It turns out they had most of the details worked out.

• Each strand of DNA is said to be complementary to the other side.

• You can tell what the other strand is made of by just looking at one strand.

Replication

• Page 298• Replication happens in the S phase of the cell

cycle. • Must happen before a cell divides in mitosis or

meiosis.• Each strand of DNA serves as a template for

the new strand that is made.• You produce one new, double stranded DNA.

Replication

• In prokaryotes, it begins at a single point on the chromosome and works both ways, up and down.

• In eukaryotes, it starts in hundreds of spots and moves until the whole thing is copied.

• It goes until the entire chromosome is replicated.

• Where replication begins is called the replication fork.

5 prime and 3 prime

• The phosphate end of the nucleotide is the 5 prime end.

• It is on the number 5 carbon.• The sugar end of the nucleotide is the 3 prime

end. • It is only the 3rd carbon.

Start of replication

• DNA is held together by two things:• Hydrogen bonds• Hydrophobic interactions-polarity• DNA helicase-uses ATP to unwind DNA.• The DNA strand that is copied is called the

template strand.• The strand that is new is called the daughter

strand.

Replication

• DNA polymerase does the work of adding bases that match up with the template strand.

• This is called semi-conservative replication.• DNA polymerase in humans punches in 50

bases a second.• There are 80 million base pairs.• Completes it in an hour.• Many, many replication forks.

DNA polymerase

• DNA polymerase can’t start without something called a primer.

• This is a short strand of RNA.• It works from the 5’ to 3’ end.• Only puts in a few bases.• U is substituted for T in RNA.• The RNA bases are added by primase.

DNA polymerase

• DNA polymerase adds its bases in the place of the primer and keeps going from 5’ to 3’.

• DNA polymerase can ONLY work from 5’ to 3’.• Only one strand moves in this direction.• This is called the leading strand.• The strand that is 3’ to 5’ prime is the “wrong”

direction.• It is called the lagging strand.

DNA polymerase

• DNA polymerase is forced to work backwards on the lagging strand.

• It makes short segments called Okazaki fragments.

• They are connected by DNA ligase. • Errors are fewer than 1 base in 1,000,000.• E. coli codes for 1,000 bases a second.

Repair Mechanisms

• Can be damaged by natural chemical alterations.

• Also by environmental agents.• Even though only one mistake is made per

1,000,000 bases, still about 50,000 bases are mutated.

• So many bases are copied that few mistakes still make large numbers.

• How did life continue with this many errors?

Ways to repair-proofreading

• Corrects errors in replication as polymerase works.

• This work is done by polymerase.• It happens every time a base is inserted.• This only misses about 1 in 10,000 bases.• This reduces the mistake rate to 1 in 1010

Mismatch Repair

• A group of proteins that follows DNA polymerase.

• This happens quickly.• The double stranded helix has not reformed

yet.• A faulty group of proteins in this group is a

common cause of colon cancer.

Excision Repair

• DNA can be damaged in other parts of the cell cycle, such as G1.

• This can be caused by radiation and chemicals.• Enzymes constantly inspect DNA even when it

is wound. • The defective strand is cut out.• The matching bases on the other strands are

cut out.

Excision Repair

• DNA polymerase and DNA ligase insert the correct bases.

• The cutting is sealed up.• This is a very complicated process, the DNA

has to be unwound to be cut, and the same type of process happens as in replication.

• This is not perfect, but it usually helps.

RNA synthesis-transcription

• Copies the information of a DNA sequence (a gene) into corresponding information in an RNA sequence.

• Makes a new RNA strand from the DNA.

RNA

• Intermediary between DNA and protein.• Differs from DNA in three ways:• Has only one polynucleotide strand• Sugar molecule is ribose rather than

deoxyribose• Uracil replaces thymine-uracil lacks a single

methyl group (CH3)

Central Dogma of Molecular Biology

• Proposed by Francis Crick.• How is DNA related to proteins?

• DNA codes for the production of RNA• RNA codes for the production of protein• Protein does not code for production of protein,

DNA, or RNA.• How does DNA get info from nucleus to

cytoplasm?

Types of RNA

• Messenger RNA-takes the info from DNA outside of the nucleus to the cytoplasm.

• Ribosomal RNA-proteins are made by ribosomes, and RNA is found on ribosomes.

• Transfer RNA-matches up the correct amino acid with the correct sequence on mRNA.

Transcription

• Requires RNA polymerase• Takes three steps:• Initiation• Elongation• Termination• RNA polymerase does not require a primer.• Still uses a template strand of DNA, the non-

template strand (coding strand) is not copied.

Transcription-Initiation

• Starts with a promoter-sometimes called a promoter sequence

• RNA polymerase binds with this sequence in DNA.

• There is at least one promoter for each gene.• Promoters tell RNA polymerase 3 things:– Where to start transcription– Which strand to transcribe– The direction to take from the start

Transcription-Elongation

• Starts when RNAP binds to promoter.• Unwinds DNA 10 base pairs at a time • Can only add bases from 5’ to 3’• Reads the template strand in the 3’ to 5’

direction.• mRNA is antiparallel to template strand.• Do not have monitors like DNA replication.• Errors are 1 in 10,000.

Transcription-Termination

• Base sequences in DNA signal termination of mRNA.

• Sometimes a helper protein pulls the mRNA away from the DNA.

• Translation can happen immediately in prokaryotes.

• In eukaryotes, mRNA has to leave the nucleus, but first be modified.

What do we have?

• We made a strand of messenger RNA using DNA as a template.

• The mRNA can pass out of the nucleus.• It goes to the cytoplasm to code for amino

acids.

Splicing

• Most mRNA strands are too long.• Page 302 figure 12-15• Contains introns and exons• Introns leave and are “spliced” or cut out.• Exons stay in and code for the protein.• Cap is 5’ and tail is 3’• This all happens in the nucleus

Splicing

• Why use the energy to cut out the introns?• Not completely sure?• Many mRNA sequences can code for different

things, depending on what is spliced.• Could only play a role in evolution, as the

genes in introns were once expressed but not used anymore.

Translation

• Changing from mRNA to code for amino acids.• Codon-a three base sequence on mRNA that

codes for a specific amino acid.• There are 20 kinds of amino acids.• The sequence of amino acids makes a protein

specialize.

Translation

• Start codon-starts translation. It is AUG and codes for methionine.

• There are three stop codons: UGA, UAG, UAA.• The stop codons make translation stop and

the polypeptide is released.• How does a codon code for amino acids?

tRNA=Transfer RNA

• tRNA must read mRNA codons correctly.• tRNA must deliver the amino acids that

correspond to the mRNA codons it has read.• Functions of tRNA:– Carries an amino acid– Associates with mRNA– Interacts with ribosomes

tRNA

• Made of about 75 to 80 nucleotides.• It has a specific 3D shape.• It is in the shape of a “T”• The top of the T has the amino acid attached.• The bottom of the T has the anticodon.• Anticodon-the opposite base sequence of the

codon.

Ribosome

• This is the workbench for translation.• It is not specific to any type of protein.• Can code for all types of proteins.• It is made of two subunits that attach when

translation starts:– Small subunit– Large subunit-has its own RNA called rRNA

Initiation of Translation

• The small subunit matches up with an introductory sequence on mRNA.

• tRNA carrying the first amino acid (methionine) attaches to the mRNA.

• The codon and anticodon match up.• The large subunit binds.• The mRNA is actually fed between the subunits.• There is a binding site for tRNA on the large

subunit.

Elongation

• The next tRNA with the matching anticodon to the codon on mRNA binds to the large subunit.

• The large subunit catalyzes the peptide bonds between amino acids.

• The mRNA is moving from the 5’ to 3’ direction.

Termination

• The cycle ends when a stop codon comes through in the mRNA.

• They do not code for amino acids, rather they code for a protein release factor.

• The polypeptide is released and the subunits separate.

Speeding up the process

• As soon as enough mRNA has passed through a particular ribosome complex, it can bind to another ribosome complex.

• Basically, the same mRNA can be coding for 3 or 4 proteins at once.

• This makes many proteins in a quick form.

Mutations

Change in DNA that alters genetic information.• Somatic Mutations-happens in somatic cells. • These mutations are not passed to sexually

produced offspring.• Germ-line mutations-passed on to the offspring in

the sex cells.• Gene mutation-mutation in a single gene.• Chromosomal mutation-mutation in a whole

chromosome.

Types of mutations

• Point mutation-a type of mutation that affects only one nucleotide.

• Types of point mutations:• Substitution-a base is substituted for another.• Insertion-a base is inserted that should not be

there.• Deletion-a base is deleted from the sequence.• Which one of these is more damaging?

Frameshift mutation

• Insertions and deletions cause a frameshift mutation.

• Everything after these types of mutations is changed because of that mutation.

• This will definitely not produce the correct protein.

Chromosomal mutations

• Duplication-a segment of the chromosome is repeated.

• Deletion-a segment of the chromosome is lost.

• Inversion-segments of the chromosome are swapped.

• Translocation-a segment of one chromosome is added to another chromosome.

Prokaryotic Gene Regulation

• How are genes turned on and off?• Look at page 309.• The picture shows regulatory genes before

the transcription promoter. • This is where specific proteins can bind and

genes can be turned on or off together.• Genes that are turned on or off together are

called operons.

Operon

• In E. coli, it is called a lac operon. • This is because this gene must be expressed

for bacteria to use the sugar lactose as food.• Lactose is a carbohydrate made of galactose

and glucose.• Bacteria are single-celled, so it must transport

lactose across the cell membrane and break the bond, creating galactose and glucose.

Operon

• Genes have to be turned on by the lac operon to produce proteins that do the work.

• If lactose is the only food source, then the bacterium has to produce these proteins to eat.

• If some other sugar is available, it doesn’t have to produce these proteins.

• The lac genes are turned off by repressors and turned on by lactose.

Operon

• Look at page 310. • The RNA polymerase binds in the promoter

region (P).• Without lactose present, a repressor binds in

the operator region, preventing RNA polymerase from transcribing.

• This turns the gene off!

Operon

• If lactose is present, it can bind to the repressor and remove it from the DNA.

• This is like a lock and key.• RNA polymerase can now bind and transcribe

the gene, producing proteins that can break down lactose.

Eukaryotic Gene Regulation

• Eukaryotes do not contain operons, but more complex sequences.

• Sequence before the initiation site called the TATA box is about 30 bases long.

• It seems to help position RNA polymerase right before it starts transcribing.

• There are also promoter sequences before the TATA box telling it to bind.

Enhancers

• Enhancers are before the promoters.• Enhancers can do the following:

• Block Transcription• Attract RNA polymerase• Tell proteins to open up the DNA sequence

Why is it so complex?

• All cells contain the same DNA.• In skin cells, you need the genes that code for

every other type of cell to be turned off.• Many genes are always turned off throughout

the life cycle of a cell.• Only a fraction of genes are ever turned on in

a specific cell.