DNA Structure & PropertiesLecture 6

Lecture Objectives

Describe the experiments that first supported the hypothesis that a cell’s hereditary material is located in the nucleus.

Explain the evidence that supports the identity of DNA as hereditary material.

Identify the three subunits of DNA and describe how they are put together to construct an intact molecule.

Describe Watson and Crick’s three--dimensional model of DNA based upon Franklin’s X-ray crystallography.

The objectives of this lecture are to :

I. DNA’s Discovery & Structure

1. A History of DNA

F. Griffiths (1928)

Tried to determine what genetic material was made of.

Griffiths’ Experiment

Avery, MacCleod & McCarthy (1944)

Tried purifying the transforming principle to change R-type Pneumococcus to S-type

The Transforming Principle is DNA

Avery, Macleod, & McCarty – 1943 Attempted to identify Griffith’s “transforming

principle” Separated the dead virulent cells into fractions

The protein fraction DNA fraction

Co-injected them with the avirulent strain. When co-injected with protein fraction, the mice lived with the DNA fraction, the mice died

Result was IGNORED Most scientists believed protein was the genetic

material.

The Hershey-Chase Experiment

Hershey & Chase – 1952 Performed the definitive

experiment that showed that DNA was the genetic material.

Chargaff’s Rule

Chargaff’s rule is a rule about DNA,

Chargaff’s Rule

Once DNA was recognized as the genetic material, scientists began investigating its mechanism and structure.

Erwin Chargaff – 1950 discovered the % content of the 4 nucleotides was

the same in all tissues of the same species percentages could vary from species to species.

He also found that in all animals (Chargaff’s rule):

%G = %C%A = %T

Watson and Crick shared the 1962 Nobel Prize for Physiology and Medicine with Maurice Wilkins. Rosalind Franklin died before this date.

The Double Helix: Watson & Crick

The Double Helix: Watson & Crick

James Watson and Francis Crick – 1953 Presented a model of the structure of DNA. It was already known from chemical studies that

DNA was a polymer of nucleotide (sugar, base and phosphate) units.

X-ray crystallographic data obtained by Rosalind Franklin, combined with the previous results from Chargaff and others, were fitted together by Watson and Crick into the double helix model.

Two types of nucleic acid can be recognized:

deoxyribonucleic acid (DNA) and ribonucleic acid

(RNA).

DNA is mostly found in the nucleus where it forms the

principal substance of the chromosomal material, the

chromatin. In addition to DNA, chromatin contains

proteins, mainly histones, and little RNA.

2. Chemical Bases in DNA

In prokaryotes, DNA is present in a single

chromosome in the nucleoid.

Little DNA is also found in mitochondria and in

chloroplasts.

Many viruses are made up of DNA, mostly double

stranded, but some are single stranded.

2. Chemical Bases in DNA

3. Primary Structure: Nucleotide & Nucleoside

The addition of a pentose sugar to a base produces a nucleoside .

If the sugar is ribose, a ribonucleoside is produced; if the sugar is 2-deoxyribose, a deoxyribonucleoside is produced

Addition of phosphate group to nucleoside produces nucleoside mono-phosphate (NMP) like AMP or CMP or a nucleotide

3. Primary Structure: Mononucleotide

PURINES1. Adenine (A)

2. Guanine (G)

PYRIMIDINES3. Thymine (T)

4. Cytosine (C) T or C

3. Primary Structure: Nitrogenous Bases

A or G

3. Primary Structure: Dinucleotide

3. Primary Structure: Polynucleotide

3’-End

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’ H

N

N

NN

N

HH

HH

N

N

NN

N

HH

H

1’

OI

O=P-O-CH2IO-

O

OH

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

OH

2’3’4’

5’

NH2

N

N

OH

H

NH2

N

N

OH

H

NH2

N

N

OH

H

NH2

N

N

OH

H

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

NH2

HN

N

NO

N NH2

HN

N

NO

N

H

NH2

HN

N

NO

N NH2

HN

N

NO

N

HH

N OH

HO

N

N

OH

H3C

N OH

HO

N

N

OH

H3C

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’ N OH

HO

N

N

OH

H3C

N OH

HO

N

N

OH

H3C

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’ N OH

HO

N

N

OH

H3C

N OH

HO

N

N

OH

H3C

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

1’

OI

O=P-O-CH2IO-

O

2’3’4’

5’

5’-EndThymine

Adenine

Cytosine

Guanine

3’ 5’ Phosphodiester bond

A

3. Primary Structure: Polynucleotide

4. Secondary structure: double helical structure

The 2 strands are twisted about each other, coiled around a common axis, forming a right- handed double helix.

The hydrophilic sugar- phosphate backbone of each chain lies on the outside of the molecule. The hydrophobic nitrogenous bases project inwards from the outer sugar-phosphate framework, perpendicular to the long axis of the helix and are stacked one above the other. The stacking of bases is held by hydrophobic bonds .This helps in holding the helical structure.

The nitrogenous bases of the 2 strands meet each other near the central axis of the helix where they become connected by hydrogen bonds between the amino, or imino, hydrogen and the ketonic oxygen atoms. The hydrogen bonding between the bases helps to hold the 2 strands of the DNA together.

GuanineThymine Adenine Cytosine

4. Secondary structure: double helical structure

A nitrogen-containing ring structure called a base. The base is attached to the 1' carbon atom of the pentose. In DNA, four different bases are found:

two purines, called adenine (A) and guanine (G)

two pyrimidines, called thymine (T) and cytosine (C)

*A always pairs with T : two hydrogen bonds

*C always pairs with G : three hydrogen bonds

4. Secondary Structure: Chargaff’s Rule

The 2 strands of the double helical molecule are antiparallel, i.e., they run in opposite direction; one runs in the 5’ to 3’ direction, while the other runs in the 3’ to 5’ direction.

4. Secondary Structure: Direction of Strands

B conformation (B-DNA): The most common form of DNA. The minor groove and major groove,

are of different widths on the outside of DNA.

A-DNA: Forms under conditions of low salt and

low humidity. There can be transient shifts from B to A

form.

Z-DNA: Consists of alternating purines and

pyrimidines Found infrequently. Z-DNA is:

long and thin Left-handed, Phosphate backbone has a zig-zag

appearance.

5. DNA Conformations

6. Key Features of a DNA molecule

DNA is the carrier

of genetic

information, which

is stored in the

form of a nucleotide

sequence. DNA has

2 important

functions:

“replication” and

“transcription”.

Replication

DNA

Transcription

RNA

Translation

Protein

7. Biochemistry of DNA

8. What is Gene ? The gene, the basic

units of inheritance; it is a segment within a very long strand of DNA with specific instruction for the production of one specific protein. Genes located on chromosome on it's place or locus.

8. What is Gene ?

A gene in relation to the double helix structure of DNA and to a chromosome (right). Introns are regions often found in eukaryote genes that are removed in the splicing process (after the DNA is transcribed into RNA): only the exons encode the protein. This diagram labels a region of only 40 or so bases as a gene. In reality most genes are hundreds of times larger.