An Introduction to Studying DNA

Gene Structure and Function:

A hallmark of all living systems...they

• Reproduce……..But how?

• Individual traits determined by genes

• Cells duplicate genes before dividing

• Genes present in gametes carry genetic information

to the next generation

What do genes do?

Genes direct the

development of an new

individual

This is done by :

directing the set of

proteins that are

produced

Determines when proteins

are produced

Genetic material has two important functions:

Must be in a form that

can be copied extremely

accurately

The correct information

must be transmitted from

cell to cell and generation

to generation

Must be translated into a

living organism

DNA is the

Genetic

Material

It is a simple molecule!!!!!!

Yet it is responsible for the

extraordinary diversity of life

DNA -it’s the genetic material

Determining it structure was one of the most

important scientific achievements of 20th

century.

It is an astonishing molecule

Structure of DNA:

• Made of only six components:

• Sugar molecule….deoxyribose

• A phosphate group….(PO4 with a negative

charge)

• Four different nitrogen containing bases

» Adenine A

» Guanine G

» Cytosine C

» Thymine T

A Nucleotide—the basic unit

Has a

Sugar

Phosphate

One of the 4 bases

The sugar: Deoxyribose

• The carbon atoms are always

numbered 1 to 5

• Carbon 1 always has a base

• Carbon 5 always has a phosphate

• Carbon 3 has an OH to attach to

another nucletoide

The DNA polymer

• The linkages formed

between the deoxyribose

and phosphate is called a

phosphodiester bond

• Creates a sugar-phosphate

backbone

• 5’ and 3’ end…….

DNA IS A POLYMER

Thousands or millions of

nucleotides are strung

together

Connect by the phosphate

on C-5 and the –OH on

C-3

Water is lost

Information of DNA is stored in

Four groups called bases

They are ringed

structures with N

Contain functional groups

that can interact

The key to the transmission of information

by DNA

The information in the

characteristics of the

bases

They form stable chemical

pairs

Complimentary Base Pairs

Adenine-thymine are said

to be complementary

Cytosine-guanine are said

to be complimentary

Base pairing:

Adenine and thymine

together form a stable

chemical pair

Cytosine and guanine form a

second stable pair.

A weak chemical interaction

called a hydrogen bond

Two stranded DNA are complimentary

• Two sugar-phosphate

backbones lie side by side

• One is arranged 5’-> 3‖

• The other one is 3’->5’

• Anti-parallel to each

other

Two stranded DNA are complimentary

The order of the specific

bases on one strand is

perfectly reflected in the

order of the

complementary bases on

the other strand.

Two stranded DNA are complimentary

Knowing the sequence of

bases on one strand

allows us to determine

the base sequence on the

complementary strand

The molecule is not flat

Each of the two sugar-

phosphate backbones is

wrapped around the other

in a conformation that is

called a double helix

Bases are inside the helix

The base pairs are on the

inside of the helix like the

rungs of a ladder

DNA function: Faithful replication

The structure of DNA

suggests how DNA is

replicated.

Either of the two strands

of DNA can be used as a

template, or pattern to

reproduce the opposite

strand

Use the rules of

complementary base

pairing

Result of DNA replication

Two daughter DNA

molecules each composed

of one parental strand and

one newly synthesized

strand identical to the

parent DNA

Semi-conservative

How does DNA replication occur

• DNA is duplicated by

enzymes

• Enzymes…..

– Unzip the double helix

– Capture free

nucleotides

– Pair the correct new

nucleotide

Make the new bond of

the sugar phosphate

backbone

• Proof-read the new DNA

strand to check for errors

Central player—How this is accomplished

Enzyme called:

DNA polymerase

Makes correct base pairs

Forms the new phosphodiester bond

Few errors occur during DNA replication

Has high fidelity

Semi-conservative replication

DNA varies from one organism to another

• Entire sum of DNA in a cell is called the GENOME

• Varies from organism to organism , but is the same in

every somatic cell within an organism

• A typical cell uses DNA to make more than 2000

different kinds of proteins…the proteins made will vary

from cell to cell

Even though each cell has all the DNA to code for the entire

organism..it does not USE all of the DNA to make proteins

Sizes of Genomes

Organism Number of

genes

Size of

Genome (bp)

Haemophilus

influenza

1,749 1,830,137

Mycoplasm

genitalium

470 580,070

C. elegans 19,899 97,000,000

Homo sapiens 40,000 3,000,000,000

DNA Function: Information Transmission

It is not so obvious how

such a simple molecule

can determine the

development of all living

things

As complex and varied as a

whale or a rose

As simple as

a bacteria

To understand how

DNA elegantly

fulfills this role

It is necessary to think about what makes a whale a whale or a rose a rose…

The answer:Proteins!

Proteins carry out nearly every function

necessary for life

• Proteins…

– Duplicate the whale’s DNA

– Structural proteins form the skin, muscles, organs and

tissues

– Transport proteins carry oxygen, nutrients hormones

and other important molecules

– Protein receptors embedded in cell surfaces bind to

whale hormones to provide the message to grow and

develop

– Immune system of proteins fight infection

– Protein catalysts digest food, synthesize fat for blubber,

replicated DNA for its baby whales and every thing

else!

Proteins are the product of DNA

The information in DNA must somehow be converted

into proteins…the ―stuff‖ of all living organisms

A protein is a chain of amino acids (20 different amino

acids) that is folded and coiled into a specific 3-D

structure

The unique 3-D structure is what makes protein function

possible

Consider a receptor protein………..

• Often embedded in a membrane

• It looks like an

irregularly shaped

glob with nooks

and cranies

• The shape is ABSOLUTELY critical to the proteins

function

Molecules Bind to Active Sites

One cranny may bind a protein on the outside of the cell

(--maybe, like a growth hormone)

This hormone must

fit exactly to signal a

cell response (in this case, growth)

Other nooks or crannies

fit appropriate molecules

to communicate with other

components of the cell

Through interaction at these sites

• The receptor protein can tell the cell that the hormone

signal has arrived.– Crucial point about protein function

» A protein depends on its ability to fit

or bind to other molecules

» That ability is determined by its three dimensional structure

» Three dimensional structure is determined by the amino acid sequence of the

protein chain

How does DNA Control

Development???????????

DNA determines the

characteristics of an

organism because it …..

…………determines the

amino acid sequences of

all the proteins in that

organism!!!

How does DNA determine an amino acid

sequence?

• DNA contains a genetic

code for amino acids

• The sequence of every

three DNA bases

represents an amino acid.

• The triplet is called a

codon.

How does DNA determine an amino acid

sequence?

• The complete stretch of

DNA needed to

determine the amino acid

sequence of a single

protein is a gene.

• A gene is the unit of

heredity defined by

classical genetics

• A complete set of genes in

an organism is its genome

Three bases --coding for amino acids

The genetic code found in

the sequence of bases in

DNA…….

but DNA does not have

U (uricil)

DNA does not directly code for AA’s

The code is read by

copying stretches of DNA

into the related nucleic

acid, RNA, in a process

called transcription.

The working copy of the

gene is called messenger

RNA (mRNA)

The Central Dogma of Biology

DNA is transcribed into

RNA (DNA and RNA use the

same language)

RNA is translated into

amino acids (proteins use a

different language)

Amino acids provide the

structure to give protein

function

RNA serves several roles

Most of these RNA

molecules are used to

synthesize proteins

Others are used directly

in structures such as

ribosomes,

spliceosomes, and

tRNA

All cells use the same genetic code

The basic DNA molecule

and the genetic code are

the same in all organisms

However the

packaging of the

DNA and

its location within

the cell varies with

the type of organism

All cells use the same genetic code, but

there are differences

Prokaryotic DNA

Floats in the cytoplasm

Usually one circular

DNA molecule

Supercoiled (folded like

a twisted rubber band)

Plasmid DNA may also

be present

Easily

regulated

by

operators

Eukaryotic DNA

Packaged into

chromosomes and

contained in nucleus

Generally several

chromosomes per cell

Much larger than

prokaryotic DNA

Regulated by enhancers

and promoters

Prokaryotic DNA

Bacteria are prokaryotic

Do not contain a nucleus

DNA is floating in the

cytoplasm

Typically contains only

one, long, circular DNA

molecule (chromosome)

Supercoiled,

Prokaryotic DNA

Prokaryotic DNA – It’s Genomic DNA

E. coli is a typical

prokaryotic organism

It has a single

chromosome with ~4,000

genes (codes for protein

and RNA)

4.6 million bp—

It is double stranded

Helical

Made of A-T/G-C base pairs

Prokaryotic DNA and Extra Chromosomal DNA

Bacteria can acquire

foreign DNA

DNA from another source

provides new characteristics to

the bacteria

Transformation

Transduction

Conjugation

Free floating DNA is

taken in by bacteria

DNA is introduced into

bacteria by a virus

Two bacteria exchange

DNA by physical contact

Prokaryotic DNA and Extra-chromosomal DNA

Bacteria often contain extra DNA

Small rings of DNA that float

in the cytoplasm called

plasmids

Plasmids contain only a few

genes (5 to 10)

Genes needed for extreme

conditions………..LIKE????

Prokaryotic DNA

Plasmids:

Most important are R

plasmids

Provide genes for

antibiotic resistance

Used in cloning—provides

a selective tool

Prokaryotic DNA

Plasmid MapPlasmids are engineered with

desired characteristics

Bacteria can transfer

plasmids……

This means they can

transfer genetic

information

Important!!!!!

Prokaryotic DNA

Plasmid Map Transformation of Bacteria

Mechanism for bacterial

evolution

Develop dangerous

antibiotic resistance

Used to transfer genes of

interest

Prokaryotic DNA

Plasmids serves as

recombinant DNA vectors

DNA of interest (a gene)

can be prepared and

inserted into a plasmid

vector

The recombinant vector is

then introduced into a

bacteria (HOW?)

Bacteria will make protein of

interest or more plasmids

Prokaryotic DNA—FOR GENE EXPRESSION

DNA

mRNA

Protein

A cell---no matter what

kind of cell—does not

need all of its proteins all

of the time…..

Cells need mechanisms

for controlling what genes

are expressed given the

circumstances in which it

exists

Prokaryotic DNA – Gene Expression

Regulation of gene expression has few controls

Often controlled as a set of genes all involved in a particular cell function.

Inducible genes vshousekeeping genes

Called operons Gene with control elements

Tryptophan operon, lacoperon and many others

Prokaryotic DNA…An operon contains

One or more genes and controlling elements

Structural genes Genes that code for proteins

Promoters/Operators Turn on or shut off gene expression

Best known operon --genes for lactose metabolism --are

inducible

Preferred sugar is glucose, but if lactose is present it can

be used……..

Prokaryotic DNA Recall the mechanism by which

proteins are made

DNA

mRNA—made by RNA polymerase

An enzyme that must attach to DNA to transcribe the DNA into RNA

Protein

RNA polymerase attaches to DNA at a specific nucleotide sequence called a promotor

TATAAT

Binding of RNA polymerase ―TURNS ON‖ a gene!

Turning on a Gene

Prokaryotic DNA Expression is Regulated

Lac operon contains

Operator---Where a

repressor sits to prevent

RNA polymerase from

working

Also a gene for repressor

Has a site for RNA

Polymerase--promoter

Three genes involved in

lactose metabolism

1 2 3Promoter Operator

Turning on a gene………

Prokaryotic DNA-Expression is regulated

E. Coli prefers to use

glucose.

The repressor blocks the

promoter when glucose is

present

This prevents RNA

polymerase from

transcribing DNA

Turning on a gene………

Prokaryotic DNA-Expression is regulated

E. Coli prefers to use

glucose….but what if

glucose is not available??

What if the sugar available

is lactose

Lactose

Turning on a gene………

Prokaryotic DNA-Expression is regulated

Lactose binds to the

repressor protein

Turning on a gene………

Prokaryotic DNA-Expression is regulated

The repressor leaves

DNA

Turning on a gene………

Prokaryotic DNA-Expression is regulated

The repressor leaves

DNA……………………

…….

RNA polymerase binds to

DNA..Makes the mRNA

for 3 genes used for

lactose

Prokaryotic DNA

When lactose is present it

can be used as a sugar

source

Must be split into glucose

and galactose

The enzyme B-

galactosidase splits the

disaccharide into two

sugars

Prokaryotic DNA

Lactose is present it binds

to the repressor protein

which falls off of DNA

RNA polymerase then

transcribes DNA into

mRNA

Ribosomes use the mRNA

to make the proteins

needed to use lactose

Prokaryotic DNA

Lac operon on U-tube

Lac operon

Lac operon 2

http://www.youtube.com/

watch?v=oBwtxdI1zvk&fea

ture=related

http://www.youtube.com/

watch?v=NfeUT3AUJd0&

NR=1

Prokaryotic DNA

Using bacteria to produce

proteins of interest

requires using

promoters

inducible systems.

Genetic control

uses promoters and

operators

turn on and off the

production of selected

genes

Eukaryotic DNA

DNA is packaged into

several chromosomes per

cell

Each chromosome is a single

linear molecule of DNA

highly coiled around histone

proteins

Each chromosome may

contain several million NT

and many thousand genes

Eukaryotic DNA

Every cell has the same number of chromosome

Humans have 46 chromosomes with 3 billion bp.

Much of the DNA is noncoding

Spacer DNA provides places for recombination

Eukaryotic DNA--REGULATION

Promoters and enhancers

provide regulation

The mRNA product must

be processed

Promoters = Sequences of

DNA where proteins bind

There are changes that

take place between the

primary transcript and the

mRNA that is translated

into a protein

Eukaryotic DNA

Promoter Exon

Proteins bind here to regulate transcription

Transcriptional factors and RNA polymerase

Intron Exon

Intervening sequence…must be cut from

mRNA

Eukaryotic DNA

Promoter Exon

mRNA

Intron Exon

RNA is processed..Introns removed…..exons spliced together

Eukaryotic DNA

DNA is expressed at a

low level without

operators

Enhancer molecules

interact with RNA

polymerase to increase

gene transcription

Genes also have

promoters

Eukaryotic DNA

Large transcriptional

complex finds structural

gene of DNA

The transcriptional

complex includes

molecules that turn on

genes

Called transcriptional

factors

Eukaryotic DNA

Only one side of DNA is

copies when making a

mRNA molecule

The mRNA is not ready

for translation

mRNA must be processed

because structural genes

contain intervening

sequences

Eukaryotic DNA

The sequences of DNA

that intervene between

the expressed sequences

are called introns.

Expressed sequences are

called exons.

Introns are sliced out of

the mRNA before it is

translated to protein

http://www.dnalc.org/reso

urces/3d/TranscriptionBasi

c_withFX.html

Eukaryotic DNA

In Biotechnology, eukaryotic DNA is used as a source of genes of interest.

Once identified, the gene can be cut from the genome or amplified (used to make many copies of the sequence).

Then used for cloning

Viral DNA

Viruses infect organism and are made of a protein coat and nucleic acid

Do not have cellular structure and are dependent upon a host cell

Once in a cell, viral DNA/RNA is released and read by host cells enzymes

Viral DNA

New viruses can be

assembled and released to

infect other cells.

Some viruses incorporate

the DNA into the host

chromosome.

Viral particles can be used

to deliver DNA , especially

to mammalian cells.

Viral DNA

Recombinant Virus

Technology is one

technique used in gene

therapy…….the

technology used to

correct defective genes in

mammals.