DNA : The Structure of chromosomes and genes

1. DNA and RNA perform different functions

What is DNA ?
• DNAs are extremely long molecules
• DNAs consisting of deoxyribonucleotides
• DNAs are double-stranded

DNA – in prokaryotic cells

The chromosome of prokaryotic cell is a single large DNA molecule, tightly bunched into a
nuclear zone or nucleoid (Prokaryotes do not possess a membrane surrounding the genetic
material)

DNA – in eukaryotic cells

• Eukaryotic cells contain many DNA molecules, each generally much larger than the
single DNA molecule in prokaryotes.
• DNA molecules in eukaryotes are combined with protein and organized into chromatin
fiber in the nucleus, which is surrounded by a complex double-membrane system.

DNA – in virus
• Virus contains nucleic acid as their genetic material, some contain DNA and some RNA
• Viral nucleic acid code for the characteristic proteins found in viral particles also for
certain enzymes required for replication of the virus in its host cell.
DNA functions
• To store the complete genetic information required to specify the structure of all
proteins and RNAs of each
• To program in time and space the orderly biosynthesis of cell and tissue component
• To determine the activities of an organism
 • To determine the individuality of a given organisms
What is RNA?
• Ribonucleic acid consist of long string of ribonucleotides
• Three major classes of RNA are messenger RNA (mRNA), Transfer RNA (tRNA), and
ribosomal RNA (rRNA)
Messenger RNA (mRNA)
• Functions as a template
• Template used by ribosomes for the translation of the genetic information into the
amino acid sequences of proteins.

Transfer RNA (tRNA)

• Consist of a single strand of ribonucleotides
• Each tRNA contains a specific trinucleotide sequence called its anticodon,
complementary to a codon

Ribosomal RNA (rRNA)

• The major component of ribosomes and constitute up to 65% of their weight
• There are three kinds of rRNA in prokaryotic and four kinds in eukaryotic ribosomes.
• rRNA play an important role in the structure and biosynthetic function of ribosomes.
There are two additional kinds of RNAs in eukaryotic cells :
1. Heterogeneous nuclear RNAs (hnRNAs), nuclear precursors of mRNAs
2. Small nuclear RNAs (snRNAs), participate in RNA processing
Small nuclear RNAs aid in removing intron RNA
2. The nucleotide units of DNA and RNA have characteristic bases and
pentoses
The Nucleotide Units of DNA and RNA
1. Nitrogenous base
2. A pentose
3. Phosphoric acids linked

The principal pyrimidine and purine bases and pentoses of DNA and RNA
 The structure of the parent compounds pyrimidine and purine

The deoxyribonucleotide units of DNA in free form at pH 7.0

DNA also contains some minor bases :
• Methylated form of the principal bases
• Hydroxymethylated or glucosylated in some viral DNA.
• Also found in RNAs, especially in tRNAs
3. Phospho-diester bonds link the successive nucleotides on nucleic acid
Phosphodiester bonds link
 • The successive nucleotides of both DNA and RNA are covalently linked to each other
through phosphate-group bridge
• Phosphate and pentose group of nucleic acid (DNA and RNA) are highly polar, since the
phosphate group are acidic and have negative charge at the pH of the cell wall
• Purine and pirimidine bases are hydrophobic

The covalent backbone structures of DNA and RNA

• The internucleotide linkage both of DNA and RNA can be chemically cleaved by
hydrolysis and enzymatically by nucleases
Several types of nuclease and their function :
• Endonuclease – hydrolyze linkage between two adjacent nucleotides at internal
positions in DNA or RNA strand
• Exonuclease – hydrolyze only the terminal nucleotide linkage, some at the 5’ and other
at the 3’ end
• Deoxyribonuclease – specific for hydrolysis DNA
• Ribonuclease – specific for hydrolysis RNA
4. DNA store genetic information
The history of DNA
1. Friedrich Miescher (Swiss Biologist) – isolated phosphorous containing substance
called nuclein from nuclei of white blood cells or leukocytes, which consist of acidic
portion which we know to day as DNA, and a basic portion, contributed by protein
2. The Avery-Macleod-McCarty experiment (1943)
 3. Hersey-Chase experiment (1952)

5. DNAs different species have distinctive base composition

Conclusion of Erwin Chargaff and his colleagues (Columbia University, 1940)
1. DNA specimen isolated from different tissues of the same species have the same base
composition
2. The base composition of DNA varies from one species to another
3. The base composition of DNA in a given species does not change with age of the
organism, its nutritional stage, or changes in its environment.
4. A = T and G = C, the sum of the purine residues equals the sum of the pyrimidine
residues;
A+G=T+C
6. Watson and Crick postulated a double-helix model of DNA structure

7. The base-sequence of DNA constitutes a template

Sequences A, T, G and C are the symbols for coding the genetic information. The sequence
functions as a template for DNA replication. Template needed for the precise replication,
transcription, and translation of genetic information. Replication of DNA during cell division
was postulated by Watson and Crick.
Replication of DNA as Suggested by Watson and Crick

8. Double-helical DNA may undergo denaturation of unwinding

9. DNA strands from two different species can perform DNA-DNA hybrids
Single strand of DNA from different species can hybrid each other when they have
complementary segment.
10. Some physical properties of double helical DNAs reflect the proportion of G=C
versus A=T pairs
 The transition from the native duplex DNA into coiled denatures form can be detected
by an increase in the absorption of ultraviolet light or a decrease in the viscosity of the
DNA solution.
 Melting point – a characteristic denaturation temperature
 Buoyant density – can be used to calculated the portion of G ≡ C and A = T

11. Native DNA molecules are very fragile

 Native DNA molecules are very fragile
 Native DNA molecules are so large that they are not readily isolated in intact form,
since they are easily broken into fragments by mechanical shear forces (stirring,
pipetting)
12. Viral DNA molecules are relatively small
Viral DNA Molecules
• Relatively small
• Table : The DNAs of some bacterial viruses
 • + since the complete base sequence of the ΦX174 (duplex form) is known, this figure is
the exact number of base pairs

13. The chromosomes of prokaryotic cells are single, very large DNA molecules
The chromosomes of prokaryote cells (DNA in an E. coli cell)
• A single – very large molecule
• Double stranded circle
• Molecular weight of about 2600 million
• Contains about 4 million base pairs
• Has a contour length of about 1400 um = 1.4 mm (700 times the length of the cell, 2
um)
• DNA molecules very tightly bunched or coiled

14. Circular DNAs are supercoiled

 Circular viral DNAs are super-coiled or supertwisted
 Endonuclease – attack super-coiled DNA
 Topoisomerases – regulate the degree of supercoiling by introducing or removing
super-twist
15. Some bacteria also contain DNA in the form of plasmids
What is plasmids?

 Extra-chromosomal DNA
 Very small
 Carry genetic information
 Can be isolated from bacterial cells
 Important in a genetic engineering
16. Eukaryotic cells contain much more DNA than prokaryotes
 Eukaryotic cells contain much more DNA than prokaryotes

 The genetic material of eukaryote cell is subdivided into chromosomes

 Eukaryotic DNAs are linier

 Each chromosome in eukaryotes carrier a unique set of genes

 Genome

17. Eukaryotic chromosomes consist of chromatin fibers

 Chromatin

 Histones

 Nucleosomes
18. Histone are small basic protein
What is histone?

 Molecular weights of histone are between 11.000 – 21.000

 Very rich in the basic amino acids (arginine and lysine)

 Have a positive charge at pH 7

19. DNA-histone complexes form beadlike nucleosomes

Levels of chromatin structure: loops (100 kbp), fibers, nucleosomes

 • DNA –histone complexes form beadlike nucleosomes

• Nucleosomes are structural units of chromatin, largely serving a packaging function

20. Eukaryotic cells also contain cytoplasmic DNA

 Mitochondria DNA (mDNA) – DNA in the cytoplasm localized in the mitochondria

 Chloroplast of photosynthetic cells also contain DNA

21. Genes are segments of DNA that code for polypeptide chains and RNAs
Genes

 Segments of DNA that code for polypeptide chains and RNAs

 One gene – one protein, one gene one enzyme, one gene one polypeptide

 Structural genes

 Regulatory sequence

 There are many genes in a single chromosomes

22. There are many genes in a single chromosomes

23. How large are genes?
The genes that code for tRNAs are much smaller than those for polypeptide, since each
nucleotide unit of tRNA is coded by a single nucleotide of DNA
24. Bacterial DNA is protected by restriction-modification systems
 Modification methylase

 Restriction endonuclease

EcoR1 Recognition Site

25. Eukaryotic DNA contains base sequences that are repeated many times
 Highly repetitive segments

 Moderately repetitive

 Non repeated segments

 Satellite DNA – non translated segments

26. A few eukaryotic genes occur in multiple copies

Most of eukaryotic genes occur in only single copies, but a few genes occur in multiple
copies.

27. Eukaryotic DNA has many palindromes

 Polindromes - inverted repetition

 Long polindromes has potential to form cruciform loops by base pairing within each
strand

Secondary Structure
28. Many eukaryotic genes contain intervening non-transcribed sequences (intron)
 Intron – intervening sequences = non translate DNA segments in genes

 Exons – the coding segment of the genes

29. The base sequences of some DNAs have been determined

Basic approaches to determining for sequencing DNAs :

• Chain termination method by Sanger and his colleagues

• Chemical method by Maxam and Gilbert

Isolation of DNA

UV Absorbance of DNA
Fluorescent Dye for DNA

Density Gradient Centrifugation