2. mRNA
• Also called as messenger RNA
• Accounts for about 5% of the total RNA in the cell.
• It is a single-stranded RNA molecule that is complementary to one of
the DNA strands of a gene
Discovery
In 1961, French scientists François Jacob and
Jacques Monod hypothesized the existence of an intermediary
between DNA and its protein products, which they called messenger
RNA.
3. Central dogma of molecular biology
The mRNA carries the message from the DNA, which controls all of the
cellular activities in a cell. If a cell requires a certain protein to be
synthesized, the gene for this product is “turned on” and the mRNA is
synthesized through the process of transcription . The mRNA then
interacts with ribosomes and other cellular machinery to direct the
synthesis of the protein it encodes during the process of translation
4. • Molecules of mRNA are composed of relatively short, single strands
of molecules made up of adenine, cytosine, guanine and uracil bases
held together by a sugar phosphate backbone.
• Like in DNA, mRNA genetic information is in the sequence
of nucleotides, which are arranged into codons consisting of
three base pairs each. Each codon codes for a specific amino acid,
except the stop codons, which terminate protein synthesis.
• mRNA in cytoplasm exist as ribonucleoproteins (covered by proteins
called informosomes) that protect mRNA from digestion by nucleases.
5. Types of mrna
Based on the types of cell they are of two types
Eukaryotic mRNA
Prokaryotic mRNA
Eukaryotic mRNA
• In eukaryotes, mRNA is transcribed on chromosomes in the nucleus, and
after processing, is shuttled through nuclear pores and into the cytoplasm
• Inside each nucleus, a multi subunit protein called RNA polymerase II
(RNAP II) reads DNA and simultaneously fabricates a “message” or
transcript
• Pre-mRNA is synthesized from a DNA template in the cell
nucleus by transcription. Pre-mRNA comprises the bulk of heterogeneous
nuclear RNA (hnRNA).
• The hnRNA is the collective term for the unprocessed mRNA molecules in
the nucleus. It contains unique sequences and has about 10 times as many
sequences as the mature mRNA
6. • The hnRNA that is associated with proteins form the heterogenous nuclear
ribonucleoprotein (hnRNP).
• In mammalian nuclei , hnRNA is the immediate product of gene
transcription The nuclear product is heterogeneous in size(Variable) and is
very large. Molecular weight may be more than 107, while the molecular
weight of m RNA is less than 2x 106
• 75 % of hnRNA is degraded in the nucleus, only 25% is processed to
mature m RNA
• Once pre-mRNA has been completely processed, it is termed "mature
messenger RNA", or simply "messenger RNA"
• which is called messenger RNA (mRNA), in a process called transcription
• Molecules of mRNA are composed of relatively short and long lived
• Eukaryotic mRNA is monocistronic which means it contains single open
reading frame (ORF) for translating only one polypeptide chain
7. • In eukaryotes, the 5’ end of mRNA is capped with a guanosine
triphosphate nucleotide, which helps in mRNA recognition during
translation or protein synthesis. Similarly, the 3’ end of an mRNA has
a poly-A tail or multiple adenylate residues added to it, which
prevents enzymatic degradation of mRNA. Both the 5’ and 3’ end of
an mRNA imparts stability to the mRNA.
8. • Cap is followed by non coding (UTR) untranslated region and then
foyllowed by a start codon and followed by a coding regions called
exons and non coding regions exons and followed by a stop codons
and untranslated region when transcription is about to end it adds a
poly A nucleotides at 3I end of RNA with the help of an enzyme poly
A polymerase
• The existence of an mRNA molecule begins with transcription, and
ultimately ends in degradation. During its life, an mRNA molecule
may also be processed, edited, and transported prior to translation.
Eukaryotic mRNA molecules often require extensive processing and
transport
9. • The extensive processing of eukaryotic pre-mRNA that leads to the
mature mRNA is the RNA splicing, a mechanism by
which introns or outrons (non-coding regions) are removed
and exons (coding regions) are joined together
• The RNA that we get after transcription is called heterogenous RNA
(hn RNA) it should be processed to become fully matured RNA
10. Prokaryotic mRNA
• In prokaryotes mRNA is produced by the process of transcription with
the help of an enzyme called RNA polymerase
• Prokaryotes lack organelles and a well defined nuclear envelope, and
therefore mRNA translation can be coupled with
mRNA transcription in the cytoplasm
• Prokaryotic mRNA is constantly degraded by ribonucleases, enzymes
that cut RNA. For example, the half-life of mRNA in E. Coli is
approximately two minutes. Bacterial mRNAs are short-lived to allow
for flexibility in adjusting to rapidly changing environmental
conditions.
11. • In prokaryotes (organisms that lack a distinct nucleus), mRNAs
contain an exact transcribed copy of the original DNA sequence with a
terminal 5′-triphosphate group and a 3′-hydroxyl residue
• 5′ region has a (UTR) untranslated region and followed by a coding
region which has a start codon and stop codon and again followed by
a UTR region
• 5′ end of a prokaryotic mRNA it has a shine dalgarno sequence which
is rich in purine nucleotides and helps in binding of mRNA to 30s
subunit of ribosome by establishing H bonds wit pyramidine
nucleotide sequence present on the 3′ end of single mRNA (16s) of
smaller subunit
12. • mRNA in prokaryotes is polycistronic it means it carries several open
reading frames (ORFs), each of which is translated into a polypeptide
13. rRNA
• It accounts for 80% of the total RNA in the cell
• The rRNA is the component of the ribosome and are located within the in
the cytoplasm of a cell, where ribosomes are found. In all living cells, the
ribosomal RNA plays a fundamental role in the synthesis and translation of
mRNA into proteins
• Molecules of rRNA are synthesized in a specialized region of the
cell nucleus called the nucleolus, which appears as a dense area within the
nucleus and contains the genes that encode rRNA.
Discovery
Ribosomal RNA was discovered during cell fractionation experiments
investigating the role of RNA viruses in causing cancer.
14. • Ribosomes consist of two major components: the small ribosomal subunits,
which read the RNA, and the large subunits, which join amino acids to form
a polypeptide chain. Each subunit comprises one or more ribosomal RNA
(rRNA) molecules and a variety of ribosomal proteins (r-protein ).
• These subunits generally are named according to their rate of sedimentation,
measured in Svedberg units
• Ribosomal proteins are synthesized in the cytoplasm and transported to the
nucleus for subassembly in the nucleolus. The subunits are then returned to
the cytoplasm for final assembly.
15. • rRNAs combine with proteins in the cytoplasm to form ribosomes, which
act as the site of protein synthesis and has the enzymes needed for the
process.
Types of rRNA
• Ribosomal RNA organizes into two ribosomal subunits: the large
ribosomal subunit (LSU) and small ribosomal subunit (SSU). Between
these subunits, the rRNA types used to form the subunit differ.
• In the ribosomes of prokaryotes such as bacteria, the SSU contains a single
small rRNA molecule (~1500 nucleotides) while the LSU contains one
single small rRNA and a single large rRNA molecule (~3000
nucleotides).These are combined with ribosomal proteins to form
ribosomal subunits.
16. • In the ribosomes of eukaryotes such as humans, the SSU contains a single
small rRNA (~1800 nucleotides) while the LSU contains two small rRNAs
and one molecule of large rRNA (~5000 nucleotides). Eukaryotic rRNA
combine with ribosomal proteins which interact to form larger and more
polymorphic ribosomal units in comparison to prokaryotes.
16S rRNA
• In bacteria the gene that has proved to be the most informative for
investigating evolutionary relatedness is 16S rRNA it is present in all
bacteria, and a related form occurs in all cells, including those of eukaryotes.
Analysis of the 16S rRNA sequences from many organisms has revealed that
some portions of the molecule undergo rapid genetic changes, thereby
distinguishing between different species within the same genus.
17. • Phylogenic information derived from the 16s rRNA is currently used as the
main method of delineation between similar prokaryotic species by
calculating nucleotide similarity
• The 3' end of the 16S ribosomal RNA (in a ribosome) recognizes a sequence
on the 5' end of mRNA called the Shine-Dalgarno sequence. Whichis used
for the correct positioning of mRNA and ribosome during the process of
translation
• The rRNA of the ribosome also has an enzymatic activity (peptidyl
transferase) and catalyzes the formation of the peptide bonds between two
aligned amino acids during protein synthesis(and thus is an enzyme
ribozyme)
18. structure
• The primary structure of rRNA sequences can vary across organisms, base-
pairing within these sequences commonly forms stem-loop configurations.
The length and position of these rRNA stem-loops allow them to create
three-dimensional rRNA structures that are similar across species.
• Because of these configurations, rRNA can form tight and specific
interactions with ribosomal proteins to form ribosomal subunits. These
ribosomal proteins contain basic residues and aromatic residues allowing
them to form chemical interactions with their associated RNA regions, such
as stacking interactions.
• Ribosomal proteins can also cross-link to the sugar-phosphate backbone of
rRNA with binding sites that consist of basic residues .
Assembly
• Ribosomal RNA's integration and assembly into ribosomes begins with their
folding, modification, processing and assembly with ribosomal proteins to
form the two ribosomal subunits, the LSU and the SSU.
• In Prokaryotes, rRNA incorporation occurs in the cytoplasm due to the lack
of membrane-bound organelles
19. • In Eukaryotes, however, this process primarily takes place in
the nucleolus and is initiated by the synthesis of pre-RNA.
• The pre-RNA then undergoes modifications such as methylation or
pseudouridinylation before ribosome assembly factors and ribosomal
proteins assemble with the pre-RNA to form pre-ribosomal particles.
• Upon going under more maturation steps and subsequent exit from the
nucleolus into the cytoplasm, these particles combine to form the ribosomes
Fully assembled subunit with proteins and rRNA
General structure of rRNA
20. tRNA
• 10 – 15% of total cellular RNA is t-RNA.
• t-RNA (transfer RNA) is also named as S-RNA (soluble or supernatant RNA)
and adaptor RNA.
• t-RNAs are small molecules with about 74 – 95 ribonucleotides,
Sedimentation constant – 3.8S, Molecular weight – nearly 25,000 – 30,000
Dalton, t-RNAs are made up of a single stranded polynucleotide chain.
• Transfer RNAs or tRNAs are molecules that act as temporary carriers
of amino acids, bringing the appropriate amino acids to the ribosome based
on the messenger RNA (mRNA) nucleotide sequence. In this way, they act
as the intermediaries between nucleotide and amino acid sequences
21. • tRNAs are ribonucleic acids and therefore capable of forming hydrogen
bonds with mRNA. Additionally, they can also form ester linkages with
amino acids
Unique feature of tRNA
• In addition to usual N-bases (A,U,G,C) tRNA contains number of
unusual bases.
• These unusual bases are important as they protect t-RNA molecules
from dehydration by Rnase, when tRNAs are floating freely in
cytoplasm.
Hydrogen bonds
Ester bond
22. • Bases that have been modified, especially by methylation (e.g. tRNA
(guanine-N7-)-methyltransferase), occur in several positions throughout the
tRNA.
• The first anticodon base, or wobble-position, is sometimes modified to
• inosine from adenine
• queuosine guanine
• uridine-5-oxyacetic acid uracil
• 5-methylaminomethyl-2-thiouridine derived from uracil
• lysidine cytosine
Activation of tRNA / charging of tRNA / aminoacylation of tRNA
• Done with the help of enzymes called aminoacyl tRNA synthetases
• It catalyses the esterification of specific aminoacid
• This enzyme catalyzes the union of amino acid to the tRNA
23.
24. Structure of tRNA
• Primary structure- linear sequence of nucleotides
• Secondary structure-Clover leaf model
• Tertiary structure- 3-D structure of tRNA , L shape,
Helix stacking
25. Primary structure
• Linear sequence of nucleotides is 60-90 in nt long but most commonly 76
• Many modified bases, sometimes accounting for 20% of the total bases in
any one tRNA molecules
• All of them are created post transcriptionally.
Secondary structure
• Robert Holley proposed clover leaf model for the first time in 1968.
• It is a two dimensional description of the t-RNA.
26. • The acceptor stem is a 7- to 9-base pair (bp) stem made by the base pairing
of the 5'-terminal nucleotide with the 3'-terminal nucleotide (which contains
the CCA 3'-terminal group used to attach the amino acid). In general, such
3'-terminal tRNA-like structures are referred to as 'genomic tags
• The amino acid loaded onto the tRNA by aminoacyl tRNA synthetases, to
form aminoacyl-tRNA, is covalently bonded to the 3'-hydroxyl group on the
CCA tail. This sequence is important for the recognition of tRNA by
enzymes and critical in translation.In prokaryotes, the CCA sequence is
transcribed in some tRNA sequences. In most prokaryotic tRNAs and
eukaryotic tRNAs, the CCA sequence is added during processing and
therefore does not appear in the tRNA gene
27. • The D arm is a 4- to 6-bp stem ending in a loop that often
contains dihydrouridine.
• The anticodon arm is a 5-bp stem whose loop contains the anticodon.The
tRNA 5'-to-3' primary structure contains the anticodon but in reverse order,
since 3'-to-5' directionality is required to read the mRNA from 5'-to-3'.
• The T arm is a 4- to 5- bp stem containing the sequence TΨC where Ψ
is pseudouridine, a modified uridine
28. • Variable arm it has between 3 and 21 nucleotides, depending on which
amino acid the tRNA encodes.
• Between anticodon loop and TΨU loop
• This tRNA's variable arm is very short so it looks quite different from the
other arms of the molecule.
• May present or absent, it depends on species.
• The length of the variable arm is important in the recognition of the
aminoacyl tRNA synthetase for the tRNA.
• Variable arm helps is stability of tRNA
• tRNAs are called class 1 if they lack it, and class 2 if they have it.
29. Teritiary structure
• It is 3D model and L shaped
• Discovered in amino acid of phenylalanine in yeast
• The acceptor stem and T-arm forming an extended helix and the anticodon
loop and D-arm similarly making another extended helix. These two helices
align perpendicularly to each other in a way that brings the D-arm and T-arm
into close proximity while the anticodon loop and the acceptor arm are
positioned on opposite ends of the molecule.
In this image, the
3’ CCA region is in yellow,
the acceptor arm is in purple,
the variable loop in orange,
the D-arm is in red,
the T-arm in green and the
anticodon loop is in blue.
30. Stability of L-structure
• Tertiary structure of t-RNA is produced by hydrogen bonding
Between N-bases
Between N-bases and ribose- phosphate backbone
Between ribose-phosphate backbone
31. Structure and Function of RNA
mRNA rRNA tRNA
STRUCTURE
Short, unstable,
single-
stranded RNA corres
ponding to a gene
encoded within DNA
Longer, stable RNA
molecules
composing 60% of
ribosome’s mass
Short (70-90
nucleotides), stable
RNA with extensive
intramolecular base
pairing; contains an
amino acid binding
site and an mRNA
binding site
FUNCTION
Serves as
intermediary
between DNA and
protein; used by
ribosome to direct
synthesis of protein
it encodes
Ensures the proper
alignment of mRNA,
tRNA, and ribosome
during protein
synthesis;
catalyzes peptide
bond formation
between amino
acids
Carries the correct
amino acid to the
site of protein
synthesis in the
ribosome
