MICROBIAL GENETICS, MCB 302

DNA and RNA

Deoxyribonucleic acid is a polynucleotide which houses the genetic ingredient of all organisms.
Its monomer unit are called nucleotides. Each nucleotide consist of a 5-carbon sugar, the
deoxyribose, a nitrogen containing base attached to the sugar and a phosphate group.
Components of DNA
1 Nitrogenous base: This is made up of two types,
i) The purines, made up of a pyrimidine plus a five member carbon and nitrogen ring.
ii) The pyrimidines, made up of carbon and nitrogen benzene ring
2 Pentose sugars: These are either ribose, as in RNA- ribonucleic acid, or DNA,
deoxyribonucleic acid. Both are 5-carbon sugars.
3 Phosphate groups, which are normally phosphoric acid.
1. NITROGENOUS BASES
These are the
Purines: Adenine and Guanine are the purines. In structure, they appear larger than
pyrimidines. The 9-atoms that make up fused ring (called Imidazole ring), of 5 carbon and 4
nitrogen are numbered in the diagram and lie in the same plane.
Pyrimidines: These are Cytosine and Thymine. Their 6-atoms of 4 carbon and 2 nitrogen are
numbered in clockwise direction, but starting from ‘6’o clock’.
 2. PENTOSE SUGARS
Ribose Sugar: This is a five-carbon sugar that forms a cyclic ring and is numbered 1’-5’.
Deoxyribose Sugar: This 5-carbon sugar has three oxygen atoms and is numbered from 1’- 5’.
The hydroxyl groups on the 5’ and 3’ links to the phosphate group to form the DNA backbone.
As the name suggests, DNA has lost one of its hydrogen atoms, as a ‘deoxyribose’ lacks a
hydroxyl group at the 2’ position, when compared to the ribose.’

NUCLEOSIDE
This is formed when a nitrogenous base covalently bonding the C1 position of the sugar. Carbon
atoms on the sugar molecules are numbered as prime to differentiate them from the number on
the atoms of the nitrogenous bases.
The sugar in deoxynucleoside is 2’-deoxyribose while that in ribonucleoside is ribose. The
nucleoside. The nucleosides are yet without the phosphate groups and are 5 in number according
to the bases:
dA- deoxyadenosine, dG- deoxyguanine, dC- deoxycytosine and dT- deoxythymidine
NUCLEOTIDES: These are the nucleosides with the phosphate groups attached to the 3-
hydroxyl and 5-hydroxyl ends of the ribose or deoxyribose in phosphodiester bonds. Regular
units of nucleotides are the DNA backbones.

3. PHOSPHATE GROUPS: These form phosphodiester bonds with the five-carbon sugars
of the DNA or RNA molecule.
DNA DOUBLE HELIX:
DNA is a double stranded macro molecule, having two polypeptide chains connected to each
other by hydrogen bonds.
Special features of double stranded DNA
- Each strand of the DNA forms a helical/spiral structure that winds around a helix in a
right handed spiral.
- DNA has two polypeptide chains that run parallel yet opposite to each other. At one end,
there is an exposed 5’ hydroxyl group while at the other end has a 3’ hydroxyl group. So,
one strand starts with 5’ and ends with 3’ while the other strand starts with 3’ and ends
with a 5’.
- The DNA has a sugar-phosphate backbone of the two strands that wind in the helix axis,
just like is obtained in spiral staircases.
- Nitrogenous bases of each nucleotides face the inner side of the helix, and look as though
they are stacked on each other.

RNA
Ribonucleic acids are polynucleotides which are normally single stranded. They have a high
molecular weight and are essential for protein synthesis. They sometimes replace DNA as bearer
of genetic information in viruses, and are thus called RNA viruses. In RNA, the ribose appended
to the nucleotide is attached by phosphodiester bonds. Three of its nitrogenous bases are similar
to that of DNA, except uracil (used in place of thymine in DNA).
There are three commonly named RNA types:
Messenger RNA (mRNA): helps ribosomes to direct the synthesis of proteins, acting between
DNA and protein.
Transfer RNA (tRNA): These small sets of RNA help in the transport of amino acids to
ribosomes in the process of growing chains of protein molecules.
Ribosomal RNA (rRNA): This is part of the ribosome and helps to align the mRNA, tRNA and
ribosomes during protein synthesis. It also catalyzes formation of peptide bonds between amino
acids.
They all take part in biochemical processes in cells and most take part in complexly regulated
cell functions and differ from each other in function, structure and formation sites.
While mRNA helps to transfer genetic info from nucleus DNA to cytoplasmic ribosomes, where
translation is occurring, tRNA and rRNA help in effective translation.
Differences between DNA and RNA
DNA BASE PAIRING
Base pairs are made up of two complementary DNA nucleotides bases which combine to form
rungs of the DNA ladder. For the DNA to bind to each other, Erwin Chargaff came up with the
rules of base pairing, which states that:
Adenine always pairs with Thymine
Cytosine always pairs with Guanine

The reason why purines cannot with another purine and vise versa with pyrimidine is due to
space constraint, as there is not enough space for two purines to fit within the helix and too much
space for the pyrimidines to get close enough together for hydrogen bonds to form between
them.
From the rules of base pairing, it can be inferred that if one can read the sequences of nucleotides
on one strand of DNA, we can deduce the complementary sequence on the other strand. We can
also infer that the total amount of Adenine on an organism is also the number of Thymine,
likewise, Guanine and Cytosine (hence G+C content).
DNA REPLICATION
DNA replication is the process of synthesizing new identical genomes from one original DNA
molecule, it occurs in all living organisms and is the basis for biological inheritance. The two
strands of DNA serve as a template for the newly forming complementary ones.
DNA replication in prokaryotes is different from that in eukaryotes because most prokaryotes
have circular DNA chromosomes. Replication for all organisms always starts at the point called
ORIGIN e.g. ori-C for E coli. For some prokaryotes, at the replication fork, DNA helix is
unwound and individual strands are replicated. Two replication forks move outwards from origin
in two different directions until all the genome has been copied. The genome with the point of
origin is known as replicon. The replication fork moves along the circular strands forming a
Greek letter θ (theta), until the two forks meet and two separate chromosomes are released.

For few other prokaryotes like E. coli, at conjugation or viral replication of phage lambda, the
Rolling Circle Mechanisms is followed. Here, one strand is nicked and the free 3’ hydroxyl end
extends. The growing point rolls around the circular template while the 5’ end is displaced and
continues increasing. Viruses use this method to produce many copies of genome rapidly from
one initiation event.
Eukaryotic DNA is linear and longer and many replication processes must occur simultaneously
so that many copies of DNA can be produced rapidly. All new DNA must be synthesized in the
5’ to 3’ direction. First, the DNA must unwind, short stretches at a time, with the aid of enzymes
called HELICASE and ATP. The strands are then kept apart from each other by their binding
with single stranded DNA binding proteins (SSBs), with TOPOISOMERASES easing out the
tension generated by the rapid unwinding. RNA polymerase called PRIMASE synthesizes RNA
primers that are complementary to the template DNA. Then DNA POLYMERASE III, using
deoxyribonucleotide triphosphates as substrates (dATP, dGTP, dCTP and dTTP) to synthesize
complementary DNA from the 3’ end of the primer.
The leading strand receives one RNA primer and is continuously extended from the primer by
DNA polymerase while the lagging strand receives several primers and is discontinuously
extended forming OKAZAKI FRAGMENTS. DNA polymerase I (RNase H) removes the RNA
primers and fill up any resulting gaps in the 5’ to 3’ direction. Okazaki fragments are then joined
by DNA ligase. Replication stops when polymerase complex reaches termination site on DNA,
i.e., the Tus protein binds with the Ter site, as seen in E. coli.
A strong fidelity of the process is required because this complex process needs to be done
accurately all the time otherwise mutations and drastic changes could occur and endanger
species. DNA polymerase III proof-reads the replication process by double checking each stage
to ensure that the bases are properly paired to the template. If they are not, it uses its 3’ to 5’
EXONUCLEASE activity to remove incorrect bases (i.e., by back-spacing) and then uses its 5’
to 3’ polymerase activity to again try to input the correct bases.