CHAPTER 10Molecular Biology of the Gene

DNA serves as the molecular basis of heredity

The “Transforming Principle” 1928

Frederick Griffith Streptococcus pneumonia bacteria

was working to find cure for pneumonia

harmless live bacteria (“rough”) mixed with heat-killed pathogenic bacteria (“smooth”) causes fatal disease in mice

a substance passed from dead bacteria to live bacteria to change their phenotype

“Transforming Principle”

The “Transforming Principle”

Transformation = change in phenotypesomething in heat-killed bacteria could still transmit disease-causing properties

live pathogenicstrain of bacteria

live non-pathogenicstrain of bacteria

mice die mice live

heat-killed pathogenic bacteria

mix heat-killed pathogenic & non-pathogenicbacteria

mice live mice die

A. B. C. D.

DNA is the “Transforming Principle”

Avery, McCarty & MacLeod purified both DNA & proteins separately from

Streptococcus pneumonia bacteria which will transform non-pathogenic bacteria?

injected protein into bacteria no effect

injected DNA into bacteria transformed harmless bacteria into

virulent bacteria

1944

What’s theconclusion?

mice die

THE STRUCTURE OF THE GENETIC MATERIAL Experiments showed that DNA is the genetic material

The Hershey-Chase experiment showed that certain viruses reprogram host cells to produce more viruses by injecting their DNA

Confirmation of DNA

Hershey & Chaseclassic “blender” experimentworked with bacteriophage

viruses that infect bacteriagrew phage viruses in 2 media,

radioactively labeled with either 35S in their proteins 32P in their DNA

infected bacteria with labeled phages

1952 | 1969Hershey

Why useSulfurvs.Phosphorus?

Protein coat labeledwith 35S

DNA labeled with 32P

bacteriophages infectbacterial cells

T2 bacteriophagesare labeled withradioactive isotopesS vs. P

bacterial cells are agitatedto remove viral protein coats

35S radioactivityfound in the medium

32P radioactivity foundin the bacterial cells

Which radioactive marker is found inside the cell?

Which molecule carries viral genetic info?

Hershey & Chase

The Hershey-Chase Experiment

Blender experiment

Radioactive phage & bacteria in blender 35S phage

radioactive proteins stayed in supernatant therefore viral protein did NOT enter bacteria

32P phage radioactive DNA stayed in pellet therefore viral DNA did enter bacteria

Confirmed DNA is “transforming factor”

Taaa-Daaa!

Hershey & Chase

Alfred HersheyMartha Chase

1952 | 1969Hershey

DNA and RNA are polymers of nucleotides

The monomer unit of DNA and RNA is the nucleotide, containing

– Nitrogenous base– 5-carbon sugar– Phosphate group

Polynucleotide Sugar-phosphate backbone

Nucleotide Phosphategroup

Nitrogenous base(A, G, C, or T)

Sugar(deoxyribose)

DNA nucleotide

Thymine (T)

DNA has four kinds of bases, A, T, C, and G

Thymine (T) Cytosine (C) Adenine (A) Guanine (G)

Pyrimidines Purines

DNA and RNA are polymers of nucleotides

RNA has a slightly different sugar (RIBOSE) RNA has Uracil instead of Thymine

Phosphategroup

Nitrogenous base(A, G, C, or U)

Sugar(ribose)

Uracil (U)

Structure of DNA

James Watson and Francis Crick worked out the three-dimensional structure of DNA, based on work by Rosalind Franklin and Maurice Wilkens

1953 | 1962

Wilkins

Rosalind Franklin (1920-1958)

DNA Structure I

The structure of DNA consists of two polynucleotide strands wrapped around each other in a double helix

DNA Structure II

Hydrogen bonds between bases hold the strands together

Each base pairs with a complementary partner

A pairs with TG pairs with C

Paired bases

DNA structuredouble helix

2 sides like a ladder

Bases match together (A pairs with T

A : TC pairs with G

C : G

DNA Structure III

Three representations of DNA

Replication of DNA

Before a cell can divide by mitosis or meiosis, it must first make a copy of its chromosomes.

The DNA in the chromosomes is copied in a process called DNA replication.

Without DNA replication, new cells would have only half the DNA of their parents.

DNA is copied during interphase prior to mitosis and meiosis.

It is important that the new copies are exactly like the original molecules.

DNA Replication depends on specific base pairing

DNA replication follows a semiconservative model– The two DNA strands separate– Each strand is used as a pattern to produce a

complementary strand, using specific base pairing– Each new DNA helix has one old strand with one new

strand

Parental moleculeof DNA

Nucleotides

Both parental strands serveas templates

Two identical daughtermolecules of DNA

DNA REPLICATION II

Untwisting and replication of DNA

Copying DNA

Matching bases allows DNA to be easily copied

Making new DNA

Copying DNA replicationDNA starts as a double-stranded

moleculematching bases (A:T, C:G)

then it unzips…

DNA replication

DNA Helicase: enzyme responsible for uncoiling the double helix and unzipping the weak hydrogen bonds between the base pairs

DNA replication

DNA polymerase

Enzyme DNA polymerase adds new bases

DNA basesin nucleus

DNAPolymerase

Copying DNA Build daughter DNA

strand use original parent

strand as “template” add new matching

bases synthesis enzyme =

DNA polymerase

• Adding bases – can only add

nucleotides to 3 end of a growing DNA strand• need a “starter”

nucleotide to bond to

– strand only grows 53

DNAPolymerase III

DNAPolymerase III

DNAPolymerase III

DNAPolymerase III

energy

energy

energy

Replication energy

3

3

5

5

DNA replication begins at the origins of replication– DNA unwinds at the origin to produce a “bubble”

– Replication proceeds in both directions from the origin

– Replication ends when products from the bubbles merge with each other

DNA replication occurs in the 5’ 3’ direction– Replication is continuous on the 3’ 5’ template

– Replication is discontinuous on the 5’ 3’ template, forming short segments

10.5 DNA replication proceeds in two directions at many sites simultaneously

Copyright © 2009 Pearson Education, Inc.

DNA replication

• Leading strand- elongates as DNA unwinds

• Lagging strand –elongates in opposite direction . Synthesized discontinuously into small segments called Okazaki fragments

• DNA ligase (an enzyme)links these sections

Limits of DNA polymerase III can only build onto 3 end of an

existing DNA strand

Leading & Lagging strands

5

5

5

5

3

3

3

53

53 3

Leading strand

Lagging strand

Okazaki fragments

ligase

Okazaki

Leading strand continuous synthesis

Lagging strand Okazaki fragments joined by ligase

“spot welder” enzyme

DNA polymerase III

3

5

growing replication fork

New copies of DNA

Get 2 exact copies of DNA to split between new cells

DNA polymerase

DNA polymerase

Copied & Paired Up Chromosomes

centromere

2009-2010

Protein Synthesis Making Proteins

How does DNA code for cells & bodies? how are cells and bodies made from the

instructions in DNA

DNA Cells Bodies

DNA has the information to build proteins genes

DNA Proteins Cells Bodies

proteinscells

bodiesDNA gets all the glory,Proteins do all the work

cytoplasm

nucleus

Cell organization

DNADNA is in the nucleus

genes = instructions for making proteinswant to keep it there = protected

“locked in the vault”

Cell organization

Proteins chains of amino acids made by a “protein factory” in cytoplasm protein factory = ribosome

nucleus

cytoplasm

ribosome

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

buildproteins

Passing on DNA information

Need to get DNA gene information from nucleus to cytoplasm need a copy of DNA messenger RNA

nucleus

cytoplasm

ribosome

mRNA

buildproteins

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

mRNA

From nucleus to cytoplasm

DNA

transcription

nucleuscytoplasm

translation

trait

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

protein

THE FLOW OF GENETIC INFORMATION FROM DNA TO RNA TO PROTEIN

The sequence of nucleotides in DNA provides a code for constructing a protein

– Protein construction requires a conversion of a nucleotide sequence to an amino acid sequence

– Transcription rewrites the DNA code into RNA, using the same nucleotide “language”

– Translation involves switching from the nucleotide “language” to amino acid “language”

DNA -> RNA -> protein.

Protein Synthesis: 2 step process

1. TRANSCRIPTION In nucleus ~ DNA -> mRNA2. TRANSLATION

in cytoplasm ~ mRNA -> ProteinInvolves 3 types of RNA1. Messenger RNA (mRNA) =carries the blueprint for construction of a protein2. Ribosomal RNA (rRNA) = the construction site where the protein is made3. Transfer RNA (tRNA) = the truck delivering the proper amino acid to the

site at the right time

10.7 Genetic information written in codons is translated into amino acid sequences

Transfer of DNA to mRNA uses “language” of nucleotidesLetters: nitrogen bases of nucleotides

(A,T,G,C)Words: codons ~triplets of bases ( ex. AGC)Sentences: polypeptide chainThe codons in a gene specify the amino

acid sequence of a polypeptide

10.8 The genetic code is the Rosetta stone of life

– Redundant: More than one codon for some amino acids

– Unambiguous: Any codon for one amino acid does not code for any other amino acid

– Does not contain spacers or punctuation: Codons are adjacent to each other with no gaps in between

– Nearly universal

10.9 Transcription produces genetic messages in the form of RNA

In transcription, the DNA helix unzips RNA nucleotides

line up along one strand of the DNA following the base-pairing rules

The single-stranded messenger RNA peels away and the DNA strands rejoin

Transcription

Making mRNA from DNADNA strand is the

template (pattern)match bases

U : AG : C

EnzymeRNA polymerase

Matching bases of DNA & RNA

Double stranded DNA unzips

A G GGGGGT T A C A C T T T T TC C C CA A

Matching bases of DNA & RNA

Double stranded DNA unzips

A G GGGGGT T A C A C T T T T TC C C CA A

Matching bases of DNA & RNA

Match RNA bases to DNA bases on one of the DNA strands

U

A G GGGGGT T A C A C T T T T TC C C CA A

U

UU

U

U

G

G

A

A

A C CRNA polymerase

C

C

C

C

C

G

G

G

G

A

A

A

AA

Matching bases of DNA & RNA

U instead of T is matched to A

TACGCACATTTACGTACGCGGDNA

AUGCGUGUAAAUGCAUGCGCCmRNAaa

aa

aa

aa

aa

aa

aa

aaaa

aa

U C CCCCCA A U G U G A A A A AG G G GU Uribosome

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

protein

cytoplasm

nucleus

traitU C CCCCCA A U G U G A A A A AG G G GU U

ribosome

10.10 Eukaryotic RNA is processed before leaving the nucleus

Noncoding segments called introns are spliced out ( coding segment = exons)

A 5’ cap and a poly A tail are added to the ends

10.11 Transfer RNA molecules serve as interpreters during translation

In the cytoplasm, a ribosome attaches to the mRNA and translates its message into a polypeptide

The process is aided by transfer RNAs

Each tRNA molecule has a triplet anticodon on one end and an amino acid attachment site on the other

Anticodon base pairs with codon of mRNA

10.12 Ribosomes build polypeptides

Translation occurs on the surface of the ribosome

– Ribosomes have two subunits: small and large

– Ribosomal subunits come together during translation

– Ribosomes have binding sites for mRNA and tRNAs

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

protein

cytoplasm

nucleus

traitU C CCCCCA A U G U G A A A A AG G G GU U

ribosome

How does mRNA code for proteins

mRNA leaves nucleusmRNA goes to ribosomes in cytoplasmProteins built from instructions on mRNA

aa aa aa aa aa aa aa aa

How?

mRNA

U C CCCCCA A U G U G A A A A AG G G GU U

How does mRNA code for proteins?

TACGCACATTTACGTACGCGGDNA

AUGCGUGUAAAUGCAUGCGCCmRNA

Met ArgVal Asn Ala Cys Alaprotein

?

How can you code for 20 amino acids withonly 4 DNA bases (A,U,G,C)?

ribosome

aa aa aa aa aa aa aa aa

AUGCGUGUAAAUGCAUGCGCCmRNA

mRNA codes for proteins in triplets

TACGCACATTTACGTACGCGGDNA

AUGGUAAAUGCAUGCGCCmRNA

Met Arg Val Asn Ala Cys Alaprotein

?

Codon = block of 3 mRNA bases

codon

ribosome

For ALL life! strongest support for a

common origin for all life

Code has duplicates several codons for

each amino acid mutation insurance!

Start codon AUG methionine

Stop codons UGA, UAA, UAG

The mRNA code

How are the codons matched to amino acids?

TACGCACATTTACGTACGCGGDNA

AUGCGUGUAAAUGCAUGCGCCmRNA

anti-codon

codon

tRNAUAC

MetGCA

ArgCAU

Val Anti-codon = block of 3 tRNA bases

aminoacid

mRNA to protein = Translation

The working instructions mRNAThe reader ribosomeThe transporter transfer RNA (tRNA)

mRNAU C CCCCCA A U G U G A A A A AG G G GU U

aaaa

aa

tRNA

GGU

aa

tRNA

U A C

aa

tRNA

GA C

tRNA

aa

A GU

ribosome

aa

aa

aaaa

aa

aa

aa

mRNA

From gene to protein

DNA

transcription

nucleuscytoplasm

protein

translation

trait

U C CCCCCA A U G U G A A A A AG G G GU Uribosome

tRNA

aa

protein

aa

aa

aa

aa

aa

aa

aa

aaaa

aa

aa

transcription

cytoplasm

nucleus

translation

trait

From gene to protein

transcriptiontranscription

translationtranslation

proteinprotein

10.15 Review: The flow of genetic information in the cell is DNARNAprotein

The sequence of codons in DNA spells out the primary structure of a polypeptide Polypeptides form

proteins that cells and organisms use

Process of protein synthesis

DNA

Transcribed strand

Transcription

Translation

RNA

Polypeptide

Startcodon

Stopcodon

10.16 Mutations can change the meaning of genes

Mutations are changes in the DNA base sequence These are caused by errors in DNA replication or by

mutagens The change of a single DNA nucleotide causes sickle-

cell disease

Mutations Any change in DNA sequence is called a mutation. can be caused by errors in replication, transcription,

cell division, or by external agents.

If mutation occurs in gametes (sex cells) it will be passed on to offspring

may produce a new trait or it may result in a protein that does not work correctly.

the mutation results in a protein that is nonfunctional, and the embryo may not survive

In some rare cases a gene mutation may have positive effects.

Mutations

If mutation takes place in a body cell, it is not passed on to organism’s offspring Damage to a gene may impair the function of the

cell When that cell divides, the new cells also will have

the same mutation Some mutations of DNA in body cells affect genes

that control cell division. This can result in the cells growing and dividing

rapidly, producing cancer.

Types of mutations

Changes to the letters (A,C,T,G bases) in the DNApoint mutation

change to ONE letter (base) in the DNAmay cause change to protein, may not

frameshift mutationaddition of a new letter (base) in the DNA

sequencedeletion of a letter (base) in the DNAboth of these shift the DNA so it changes how the

codons are readbig changes to protein!

Mutations

Point mutationssingle base change

silent mutation no amino acid change redundancy in code

missense change amino acid

nonsense change to stop codon

Mutations

Frameshift shift in the reading

framechanges everything

“downstream” insertions

adding base(s)deletions

losing base(s)

Where would this mutation cause the most change:beginning or end of gene?

THERAATANDTHECATATETHEREDBATTHERAATANDTHECATATETHEREDBAT

Frameshift mutations

THERATANDTHECATATETHEREDBAT

THERTANDTHECATATETHEREDBAT

THERATANDTHECATATETHEREDBAT

THERTANDTHECATATETHEREDBAT

Deletion

Insertion

Causes of Mutations

sometimes a mistake in base pairing during DNA replication.

many mutations are caused by factors in the environment

Any agent that can cause a change in DNA is called a mutagen. Mutagens include radiation, chemicals, and even

high temperatures