RNA AND ITS STRUCTURE, FUNCTION AND TYPES
Ribonucleic acid (RNA) is a type of molecule that consists of a long chain of nucleotide units. Each
nucleotide consists of a nitrogenous base, a ribose sugar, and a phosphate. RNA is very similar to DNA,
but differs in a few important structural details: in the cell, RNA is usually single-stranded, while DNA is
usually double-stranded; RNA nucleotides contain ribose while DNA contains deoxyribose (a type of
ribose that lacks one oxygen atom); and RNA has the base uracil rather than thymine that is present in
DNA.
Ribosomal RNA –
rRNA Ribosomal RNA, as the name suggests, is found in the ribosomes. It comprises about 80% of the
total RNA of the cell. The base sequence of rRNA is complementary to that of the region of DNA where it
is synthesized. In eukaryotes ribosomes are formed on the nucleolus. Ribosomal RNA is formed from
only a small section of the DNA molecule, and hence there is no definite base relationship between rRNA
and DNA as a whole.
Ribosomal RNA is stable for at least two generations. The ribosome consists of proteins and RNA. The
70S ribosome of prokaryotes consists of a 30S subunit and a 50S subunit. The 30S subunit contains 16S
rRNA, while the 50S subunit contains 23S and 5S rRNA. The 80S eukaryote ribosome consists of a 40S
and a 60S subunit. In vertebrates the 40S subunit contains 18S rRNA, while the 60S subunit contains 2829S, 5.8S and 5S rRNA. In plants and invertebrates the 40S subunit contains 16- 18S RNA, while the 60S
subunit contains 25S and 58 and 5.8S rRNA.
Messenger RNA - mRNA - Jacob and Monod (1961) proposed the name messenger RNA for the RNA
carrying information for protein synthesis from the DNA (genes) to the sites of protein formation
(ribosomes). It consists of only 3 to 5% of the total cellular RNA.
Messenger RNA is always single stranded. It contains mostly the bases adenine, guanine, cytosine and
uracil. There are few unusual substituted bases. Although there is a certain amount of random coiling in
extracted mRNA, there is no base pairing. In fact base pairing in the mRNA strand destroys its biological
activity Since mRNA is transcribed on DNA (genes), its base sequence is complementary to that of the
segment of DNA on which it is transcribed. This has been demonstrated by hybridization experiments in
which artificial RNA, DNA double strands are produced. Hydrization takes place only if the DNA and RNA
strands are complementary. Usually each gene transcribes its own mRNA. Therefore, there are
approximately as many types of mRNA molecules as there are genes. There may be 1,000 to 10.000
different species of mRNA in a cell. These mRNA types differ only in the sequence of their bases and in
length. When one gene (cistron) codes for a single mRNA strand the mRNA is said to be monocistronic. In
many cases, however, several adjacent cistrons may transcribe an mRNA molecule, which is then said to
be polycistronic or polygenic.
The mRNA molecule has the following structural features:
1. Cap. At the 5' end of the mRNA molecule in most eukaryote cells and animal virus molecules is found a
'cap'. This is blocked methylated structure, m7Gpp Nmp Np or m7Gpp Nmp Nmp Np. where: N = any of
the four nucleotides and Nmp = 20 methyl ribose. The rate of protein synthesis depends upon the
presence of the cap. Without the cap mRNA molecules bind very poorly to the ribosomes.
2. Noncoding region 1 (NC1). The cap is followed by a region of 10 to 100 nucleotides. This region is rich
in A and U residues, and does not translate protein.
3. The initiation codon is AUG in both prokaryotes and eukaryotes
4. The coding region consists of about 1,500 nucleotides on the average and translates protein It is made
up of 73-93 nucleotides
Transfer RNA – tRNA
Several models of the secondary structure of tRNA have been proposed, and of these the cloverleaf
model of Holley is the most widely accepted.
Transfer RNA (tRNA) is an essential component of the protein synthesis reaction. There are at least
twenty different kinds of tRNA in the cell1 and each one serves as the carrier of a specific amino acid to
the site of translation. tRNA's are L-shaped molecules. The amino acid is attached to one end and the
other end consists of three anticodon nucleotides. The anticodon pairs with a codon in messenger RNA
(mRNA) ensuring that the correct amino acid is incorporated into the growing polypeptide chain. The Lshaped tRNA is formed from a small single-stranded RNA molecule that folds into the proper
conformation. Four different regions of double-stranded RNA are formed during the folding process.
The two ends of the molecule form the acceptor stem region where the amino acid is attached. The
anticodon is an exposed single-stranded region in a loop at the end of the anticodon arm. The two other
stem/loop structures are named after the modified nucleotides that are found in those parts of the
molecule. The D arm contains dihydrouridylate residues while the TΨC arm contains a ribothymidylate
residue (T), a pseudouridylate residue (Ψ) and a cytidylate (C) residue in that order. All tRNA's have a
similar TΨC sequence. The variable arm is variable, just as you would expect. In some tRNA's it is barely
noticable while in others it is the largest arm. tRNA's are usually drawn in the "cloverleaf" form (below)
to emphasize the base-pairs in the secondary structure.
POST TRANSCRIPTIONAL MODIFICATION OF RNA
A primary transcript is a linear copy of a trnscriptional unit, the segment of DNA between specific
initiation and termination sequences. The primary transcripts of both prokaryotic and eukaryotic tRNAs
and rRNA are post transcriptionally modified by cleavage of the original transcripts by ribonucleases.
tRNAs are then further modified to help give each species its unique identity. In contrast, prokaryotic
mRNA is generally identical to its primary transcript, whereas eukaryotic mRNA is extensively modified
post transcriptionally.
Ribosomal RNA Ribosomal RNAs of both prokaryotic and eukaryotic cells are synthesized from long
precursor molecules called preribosomal (45S)RNAs. The 23S, 16S and 5S ribosomal RNAs of prokaryotes
are produced from a single RNA precursor molecule, as are the 28S, 18S, and 5.8S rRNAs of eukaryotes.
rRNA Eukaryotic 5S rRNA is synthesized by RNA polymerase III and is modified separtely. These
precursors are cleaved to yield intermediate-sized pieces of rRNA, which are further “trimmed” to
produce the required ribosomal RNA species. (Note: some of the proteins destined to become
components of the ribosome associate with the rRNA precursor prior to and during its posttranscriptional modification in the nucleolus.) Transfer RNA Both eukaryotic and prokaryotic transfer
RNAs are also made from longer precursor molecules that must be trimmed. Other post-transcriptional
modifications include additin of a – CCA sequence by nucleotidyltransferase on the 3’ terminal end of
tRNAs, and modification of bases at specific positions to produce “unusual bases”
TRANSLATION
Translation is the production of a polypeptide (protein) using RNA as a template and tRNA molecules as
“adapters” that convert the nucleic acid code to protein code. B. The nucleotides (letters) of RNA formed
codons (words) that specify a particular amino acid. C. The tRNA contains an anticodon that is
complementary to the codon and carries a specific amino acid. D. Important elements of the genetic
code: 1. The code is a triplet code: Each mRNA codon (word) that specifies a particular amino acid in a
polypeptide chain consists of three nucleotides (letters). For example, AAG = lysine 2. The code is nonoverlapping: The mRNA encoding one protein is read in successive groups of three nucleotides. 3. The
code is degenerate: More than one mRNA codon (word) occurs for some amino acids (ie. AAG and AAA
are read as both read as lysine)
4. The code has start signals (AUG and rarely GUG) and stop signals (UAA, UAG, and UGA). Stop signals
are also called nonsense codons because they do not designate an amino acid.
5. The code is commaless.
6. The code is almost universal.
7. The code… … ..
Factors required for translation
1. mRNA – contains a RBS (ribosome binding site ) / also known as a ShineDelgarno sequence. The RBS
is characterized by a core sequence 5’AGGAGU3’ located 7+2 nucleotides from the AUG. Deviations from
the consensus decrease translation.
2. tRNA – adapter molecules in the information transfer between mRNA and protein which has: a)
anticodon which is a 3 nucleotide sequence that is complementary and antiparallel to the mRNA codon
b) amino acid attachment site at the 3’ end for attachment of the amino acid c) 3-D shape that
determines which amino acid will be attached to the amino acid attachment site. Recent studies indicate
that the anticodon loop, the D loop, and the aminoacyl stem are all important. The correct attachment of
the amino acid to its tRNA is considered the “2 nd genetic code” and is still being cracked.
3. Aminoacyl tRNA synthetase – transfers the amino acid to its proper tRNA; there are 20 of these, each
recognizing 1 amino acid and all the tRNAs that to which that amino acid is to be attached 4. Ribosomes
(Note S refers to a sedimentation value of the structure in a sucrose gradient) a) Large subunit (50S)–
consists of 23S and 5S rRNAs and 31 ribosomal proteins b) Small subunit (30S)- consists of 16S rRNA and
21 ribosomal proteins 5. Soluble transcription factors a) Initiation factors (1) IF1 – promotes dissociation
of ribosomal subunits (2) IF2(·GTP) – required for fMET-tRNAmet binding (3) IF3 – required for mRNA
binding, finding the AUG b) Elongation factors (1) EF-Tu (·GTP) – binds aminoacid-tRNA to the ribosome
(2) EF-Ts – regenerates EF-Tu·GTP (3) EF-G(·GTP) – increases translocation rate c) Termination factors (1)
RF1 – recognizes UAA and UAG stop codons (2) RF2 – recognizes UAA and UGA nonsense codons (3)
RF3(·GTP) – enhanced RF-1 and –2 binding to ribosome 6. Amino acids 7. F-met (N-formyl Met-tRNA) 8.
GTP 9. ATP (for charging tRNAs)
Translation – Mechanism in Prokaryotes A. Initiation – the purpose of this step is to set the reading
frame
1. IF1, IF2·GTP, and IF3 bind to the 30S subunit.
2. Binding of mRNA to the 30S subunit via an interaction between the RBS on the RNA and a
complementary sequence at the 3’ end of the 16S RNA. Facilitated by IF3.
3. Release of IF3
4. fMET·tRNA binds to the P site in the 30S subunit with the help of IF-2.
5. GTP hydrolysis and release of IF1 and IF2 drives the attachment of the 50S subunit.
B. Elongation – addition of amino acids to the growing polypeptide chain
1. EF-Tu·GTP-AA-tRNA complex binds to the A site (there is a selection process that goes on at this step
whereby if the match between the codon and anticodon is not correct, the complex is released before
the next step can occur)
2. GTP hydrolysis
3. Proofreading (if the match between the codon and anticodon is not correct, the complex is released
before the next step can occur)
4. EF-Tu release (Note that EF-Tu·GTP is regenerated via the action of EF-Ts)
5. Peptidyl transfer – polypeptide is transferred from the tRNA at the P site to the AA-tRNA complex at
the A site. This catalytic activity is thought to involve not only proteins but also the 23S RNA.
6. Translocation – shift of the ribosome one codon towards the 3’ end resulting in transfer of the tRNA
with the polypeptide chain to the P site (stimulated by EFG). During translocation the uncharged tRNA in
the P site is moved to the E site (for exit) which is thought to block the A site unit translocation is
complete.
C. Termination
a) A termination codon (UAA, UAG, UGA) is presented at the A site
b) RF1 or RF2 bind to the A site with the help of RF3·GTP
c) The RF-mRNA-ribosome complex catalyzes peptidyl hydrolysis instead of transfer
d) The polypeptide is released from the tRNA in the P site.
e) The GTP associated with RF3 is hydrolyzed causing the release of the 3 RF factors and the tRNA from
the ribosome f) The 30S and 50S subunits dissociate with the aid of IF1 and IF3 and the mRNA is released
Elongtion