22.1. Nitrogenous base
They are :-
- Water insoluble
Nitrogenous bases are of two types :
Major pyrimidenes which are commonly found in DNA are thymine (T) and Cytosine (C) cytonine (C) and Uracil (U) are found in RNA. Pyrimidines are derived from the single ring structure known as pyrimidine ring (2N+4C).
Major purines which are commonly found in DNA and RNA are Adenine (A) and Guanine (G). Purines are derived from the two ring structure known as purine ring. One rings is pyrimidine (2N+4C) and another is imidazole ring (2N+3C).
Adenine has –NH2 group at C6 position. Guanine has –NH2 group at C2 position and carbonyl group at C6 position.
Sugars : Sugars are of two types :
- Deoxy ribose Sugar
- Ribose sugar
Deoxy Ribose Sugar :
This pentose sugar is found is DNA. In this sugar ‘H’ is present at the 2¢ position. So this sugar know as 2'–deoxy–D–ribose sugar.
Ribose Sugar :
This pentose sugar is found in RNA. In this sugar ‘OH’ group is present at the 2' position at the place of ‘H’. Due to presence of ‘OH’ group at 2'-3' this sugar also known as D-ribose sugar.
Three phosphate groups are present at the position C5. These phosphate group know as a, b, g for the first, second and third phosphate respectively.
-glycosidic bond is formed between 9 nitrogen of Adenine and Guanine and the 1' OH group of is known ribose.
In the pyrimidines b-glycosidic bond form between first nitrogen of their ring at the place of ninth nitrogen and the 1'OH group of deoxy ribose. The sugar and base alone know as nucleoside. Sugar, nitrogenous base and phosphate to gather called as nucleotide. Addition of a phosphate group to a nucleoside forms a nucleotide.
22.2. Phospho diester bond :
Many nucleotides are joined to each other through the 3' OH of 2' deoxy ribose of one nucleotide and the phosphate attached to the 5' OH group of another nucleotide. This linkage between 3' OH and 5' P are known as phospho diester bond. Condensation of two or more nucleotides from polynucleotide.
22.3. Types of DNA structure
- Primary structure : The linear sequence of nucleotide that are linked by phospho diester bond are known as primary structure of nucleic acid. Primary structure of nucleic acid consist Nitrogen base (A, G, C, T), pentose sugar and one or more phosphate groups.
- Secondary structure : Formation of hydrogen bonds between two complementary bases of double helix DNA is known as second structure of DNA. Secondary structure is responsible for the shape of DNA double ring structure purine always from hydrogen bond with single ring pyrimidine bases.
Adenine forms two Hydrogen bonds with thymine and guanine and three Hydrogen bonds with cytosine.
[A = T, G º C]
- Tertiary structure : Tertiary structure of the DNA is more condensed than the secondary structure of DNA, in which entire chain of polynucleotide is folded into a specific three dimentional shape. In tertiary structure hydrogen bonding and Hydrophobic force, (base stacking force) play an important role.
- Quaternary Structure :
It is the higher level of organization of DNA. In this structure, DNA interacts with other molecules chromatin is the quaternary structure of DNA in which it interact with histone proteins.
22.4 Different conformation of DNA :
Conformation of DNA depends on the hydrogon level i.e. (if concentration of water is high, it will be, B form of DNA, and if concentration. of water is low, it will be, A form of DNA), DNA sequence, concentration of salt and metal ions in solution and modification of the nitrogenous bases.
22.4.1 B-DNA :
It is the most abundant form of DNA in cells under normal physical condition. This form of DNA observed at the high water concentration (high humidity).
B form of DNA have following characteristics :-
- Right handed helix rotation sense.
- More twisted.
- One strand 5' 3' and other strand 3' 5' direction oriented.
- Diameter of helix is 20Å
- Distance between two adjacent basepair is 0.34 Å
- 10.5 base pair per helical turn.
22.4.2. A-DNA :
The ‘A’ form of DNA is observed under low humidity (low water concentration) condition. B form DNA adopt the ‘A’ form structure in some DNA-protein complex. ‘A’ form of DNA have following characteristics :
- Right handed
- 11 bp per turn
- Major groove is narrow and much deeper.
- Minor groove is broader and shallower.
- Distance between two adjacent base pair is 2.3Å.
- 'A' form of DNA is similar to the double helical RNA.
22.4.3. Z-DNA :
It is the left handed DNA. In the B-DNA glycosidic bond are anti conformation at pyrimidine and purine residue. But in the Z-DNA anti conformation is out only at the pyrimidine residue and syn conformation at purine residue. This syn conformation is responsible for left handedness of helix. The alternating anti-syn conformation gives the back bone of left handed DNA a zig-zag look. So this form of DNA is known as zig-zig or ‘Z’ form.
‘Z’ form of DNA have following characteristics
- Super helical tension.
- More narrow and more elongated helix.
- Major groove is not really a groove.
- Occurs in the high salt concentration.
22.5. Nucleic acid stability
Many forces stabilizing the DNA structure
- Base pairing : Base pairing holds both of the strands of DNA together like ‘glue’. Nitrogenous base of a strand form the pairs with another strand. Watson crick base pairing is the more stable of all base pairing in the double helix. It plays a crucial role roll in stability of DNA.
- Hydrophobic force : It is a tendency of water molecules to form a bigger cage around the hydrophobic molecules. In the DNA, pyrimidines and purines are hydrophobic, so water molecules forms the hydrophopic interaction with nitrogenous bases. Hydrophobic force plays a central role in determining nucleic acid structure.
- Base stacking interaction : Interaction between two adjacent nitrogenous bases which are out in plane, in the parallel manner. Different sets of base pairs in a stack have different stacking energies. Stacked G and C bases have greater interaction compare to stacked A and T bases. This force helps to minimize the contact of water with nitrogenous bases.
- Hydrogen bond : When a hydrogen atom present between two electronegative molecules than it forms an extra bond. This extra bond is known as hydrogen bond. Adenine is always hydrogen-bonded to thymine and guanine is always hydrogen bonded to cytosine by watson-crick pairing. The surface atoms in the sugar-fourth and phosphate form hydrogen bond to water molecules.
- Ionic interaction : Phosphate group of back bone of DNA play an important role in DNA stability due to their one charge. If we add the positive ions (Na+) in the DNA solution, the melting temperature of duplex DNA increase means stability of DNA increase because these ion shield the anionic phosphate group and decrease the repulsion.
22.6. Thermal Stability
The temperature at which 50% DNA in given sample become denatured, known as melting temperature (Tm). The double stranded DNA show the less absorbance at 260 nm. While the denature DNA show the more absorbance at 260 nm this phenomena known as hyperchromatic effect.
Factors affecting Tm
- Ionic strength : When the lower the ionic strength is low it can not completely shield the anionic phosphate. So repulsion makes it energetically more favorable to separate the strands thus Tm temperature become lower.
- AT and GC Content : GC content has higher number of Hydrogen bond and base stacking interaction compare to AT content. So the high GC content has higher Tm than DNA with high AT content.
- pH : If the pH is greater than 10, the hydrogen bonds of Nitrogenous bases are destroyed and DNA become denatured. If the pH is less than 3, it also disrupt, the base pairing due to extensive protonation. So very high or very low pH lowers the Tm.