NUCLEIC ACIDS

Topic Outline:

History of Nucleic Acids Structure and Function Types of Nucleic Acids

1. DNA2. RNA

Central Dogma of Life

Friedrich Miescher in 1869

isolated what he called nuclein from the nuclei of pus cells

Richard Altmann in 1889

Nuclein was shown to have acidic properties, hence it became called nucleic acid

1920s

the tetranucleotide hypothesis was introduced

The Tetranucleotide hypothesis

Up to 1940 researchers were convinced that hydrolysis of nucleic acids yielded the four bases in equal amounts.

Nucleic acid was postulated to contain one of each of the four nucleotides, the tetranucleotide hypothesis.

Takahashi (1932) proposed a structure of nucleotide bases connected by phosphodiester linkages.

The Tetranucleotide hypothesis

adenine uracil

cytosine guaninephosphate

phosphate phosphate

phosphate

pentose

pentosepentose

pentose

Astbury and Bell in 1938

First X-ray diffraction pattern of DNA is published.

The pattern indicates a helical structure, indicated periodicity.

X-ray diffraction of DNA

Wilkins & Franklin (1952): X-ray crystallography

Avery, MacLeod, and Mc Carty in 1944

demonstrate DNA could “transform” cells.

Supporters of the tetranucleotide hypothesis did not believe nucleic acid was variable enough to be a molecule of heredity and store genetic information.

Mice injected with virulent bacteria die

The Avery, MacLeod, and McCarty Experiment

Mice injected with nonvirulent bacteria live

The Avery, MacLeod, and McCarty Experiment

Mice injected with heat-killed virulent bacteria live

The Avery, MacLeod, and McCarty Experiment

Mice injected with a mixture of nonvirulent bacteria and heat-killed virulent bacteria die

The Avery, MacLeod, and McCarty Experiment

Mice injected with a mixture of nonvirulent bacteria and DNA from heat-killed virulent

bacteria die

The Avery, MacLeod, and McCarty Experiment

Erwin Chargaff in late 1940s

used paper chromatography forseparation of DNA

hydrolysates. Amount of adenine is equal to

amount of thymine and amount of guanine is equal to amount of cytosine.

Hershey and Chase in 1952

confirm DNA is a molecule of heredity.

The Hershey-Chase Experiment

The Hershey-Chase Experiment

Watson and Crick in 1953

determine the structure of DNA

Watson & Crick Base pairing

Francis Crick in 1958

proposes the “central dogma of molecular biology” .

Kornberg purifies DNA polymerase I

1969

Entire genetic code determined

Important Developments 1972 – First recombinant DNA molecule

constructed Lambda phage DNA inserted into SV40 virus

- Restriction enzyme EcoRI used to

create a recombinant plasmid containing penicillin and tetracycline resistance

1982 – Tetrahymena ribosomal RNA splicing shown to be self- splicing

Important Developments

1986 – RNA is shown to act as an enzyme

- The polymerase chain reaction

(PCR) was developed by Kari Mullis at Cetus

Corporation. 1995 – First complete genome

sequenced of the bacteriumHaemophilus influenzae at TIGR

2001 – Human genome sequenced by

NIH and Celera Genomics

Nucleic Acids

• Nucleic Acids are very long, thread-like polymers, made up of a linear array of monomers called nucleotides.

• Nucleic acids vary in size in nature

• tRNA molecules contain as few as 80 nucleotides

• Eukaryotic chromosomes contain as many as 100,000,000 nucleotides.

Two types of nucleic acid are found

Deoxyribonucleic acid (DNA) Ribonucleic acid (RNA)

DNA and RNA

DNAdeoxyribonucleic acidnucleic acid that stores genetic informationfound in the nucleus of a mammalian cell.

RNAribonucleic acid3 types of RNA in a cellRibosomal RNAs (rRNA) are components of ribosomesMessenger RNAs (mRNA) carry genetic informationTransfer RNAs (tRNA) are adapter molecules in translation

The distribution of nucleic acids in the eukaryotic cell DNA is found in the nucleus

with small amounts in mitochondria and chloroplasts

RNA is found throughout the cell

The nucleus contains the cell’s DNA (genome)

Nucleus

RNA is synthesized in the nucleus andexported to the cytoplasm

Nucleus

Cytoplasm

dA dG dT dC

Deoxyribonucleotides found in DNA

A G U C

Ribonucleotides found in RNA

DNA as genetic material: The circumstantial evidence1. Present in all cells and virtually restricted to the

nucleus

2. The amount of DNA in somatic cells (body cells) of any given species is constant (like the number of chromosomes)

3. The DNA content of gametes (sex cells) is half that of somatic cells. In cases of polyploidy (multiple sets of chromosomes) the DNA content increases by a proportional factor

4. The mutagenic effect of UV light peaks at 253.7nm. The peak for the absorption of UV light by DNA

NUCLEIC ACID STRUCTURE

Nucleic acids are polynucleotides

Their building blocks are nucleotides

NUCLEOTIDE STRUCTURE

PHOSPATE SUGAR

Ribose or Deoxyribose

NUCLEOTIDE

BASEPURINES PYRIMIDINES

Adenine (A)Guanine(G)

Cytocine (C)Thymine (T)Uracil (U)

All nucleotides contain three components:1. A nitrogen heterocyclic base2. A pentose sugar3. A phosphate residue

Nucleotide Structure

Ribose is a pentose

C1

C5

C4

C3 C2

O

RIBOSE DEOXYRIBOSE

CH2OH

H

OH

C

C

OH OH

C

O

H HH

C

CH2OH

H

OH

C

C

OH H

C

O

H HH

C

Spot the difference

Ribonucleotides have a 2’-OHDeoxyribonucleotides have a 2’-H

Chemical Structure of DNA vs RNA

THE SUGAR-PHOSPHATE BACKBONE

The nucleotides are all orientated in the same direction

The phosphate group joins the 3rd Carbon of one sugar to the 5th Carbon of the next in line.

P

P

P

P

P

P

ADDING IN THE BASES

The bases are attached to the 1st Carbon

Their order is important It determines the genetic information of the molecule

P

P

P

P

P

P

G

C

C

A

T

T

DNA IS MADE OF TWO STRANDS OF POLYNUCLEOTIDE

P

P

P

P

P

P

C

G

G

T

A

A

P

P

P

P

P

P

G

C

C

A

T

T

Hydrogen bonds

DNA IS MADE OF TWO STRANDS OF POLYNUCLEOTIDE

The sister strands of the DNA molecule run in opposite directions (antiparallel)

They are joined by the bases Each base is paired with a specific partner:

A is always paired with T G is always paired with C“Purine with Pyrimidine”

The sister strands are complementary but not identical

The bases are joined by hydrogen bonds, individually weak but collectively strong

There are 10 base pairs per turn

Structure of Nucleotide Bases

Purines & Pyrimidines

5’ End

3’ End

Nucleotides arelinked byphosphodiesterbonds

Three-dimensional structure of B DNA

NMR solution structure of a 12 nucleotide fragment of DNA

5’- C G C G A A T T C G C G –3’

3’- G C G C T T A A G C G C –5’

Total of 32 hydrogen bonds.

Reading Assignment: Tjandra et alJournal of the American Chemical Society (2000) 122, 6190

Cellular Processes

DNA

RNA (mRNA)

Proteins

replication

transcription

translation

RNA and Transcription

DNA is in the nucleus

Proteins are synthesized on ribosomes in the cytoplasm

RNA carries the genetic information from the nucleus to the cytoplasm

This RNA is called messenger RNA (mRNA)

Transcription of a DNA molecule results in a mRNAmolecule that is single-stranded.

RNA molecules do not have a regular structure like DNA.

The structures of RNA molecules are complex and unique.

RNA molecules can base pair with complementary DNAor RNA sequences.

G pairs with C, A pairs with U, and G pairs with U.

RNA Structure

RNA structures have many hairpins and loops

Structure of RNA Hairpin