RNA
There are
three major types of RNA that participate in the process of protein synthesis:
rRNA, tRNA, and mRNA.
First:
rRNA:
a) Are found in association with several
proteins as components of the ribosomes, the complex structures that serve as
the sites for protein synthesis.
b) There are three distinct size species of rRNA
(23S, 16S, and 5S) in prokaryotic cells while in eukaryotes there are 4
species of rRNA (28S, 18S, 5.8S, and 5S).
c) Make up about 80% of the total RNA in the cell.
Second:
tRNA:
a) Is the smallest (4S) of the three
major types of RNA molecules.
b) There is at least one specific type of tRNA
molecule for each of the 20 amino acids commonly found in proteins.
c) Make up about 15% of the total RNA in the cell.
d) Contain a high percentage of unusual
bases (Pseudouridine), for example, dihydrouracil and have extensive
intrachain base-pairing.
e) Serves as an “adaptor” molecule that
carries its specific amino acid, covalently attached to its 3ʹ-end, to the site
of protein synthesis. (amino acid is attached to terminal adenine
ribonucleotide at either 2’
or 3’ (OH) group.
Third:
mRNA:
a) Comprises only about 5% of the
RNA in the cell.
b) Carries genetic information from DNA for
use in protein synthesis.
c) Has special structural characteristics: a
long sequence of adenine nucleotides (a poly-A “tail”) on the 3ʹ-end of
the RNA chain, plus a “cap” on the 5ʹ-end consisting of a molecule of 7-methylguanosine
attached through an unusual (5ʹ→5ʹ) triphosphate linkage.
d) In eukaryotes, it undergoes
changes post transcription to produce mature mRNA or heterogenous
nuclear mRNA where introns which are non-coding regions are spliced and exons
which are coding regions are bound together to form mature one.
Transcription
process in prokaryotes:
RNA
polymerase is the
enzyme that is responsible for transcribing a part of DNA into mRNA in
direction from 5’ to 3’
of newly forming strand.
Before we
talk about the process, we will illustrate the structure of RNA polymerase.
RNA
polymerase structure:
a) Core enzyme: 2α, 1β, 1βʹ, and1Ω are part of enzyme peptide subunits where its function
is 5’ to 3’
RNA polymerase activity and have no 3’
to 5’ exonuclease (proofreading
capabilities).
b) Holoenzyme: where, the s subunit (sigma
factor) enables RNA pol to recognize promoter regions on DNA. The s
subunit with core enzyme forms holoenzyme.
Before we
illustrate steps of process, we have to explain some terms related to
transcription:
1) Sense
strand
is the DNA strand that is similar to mRNA.
And it is in direction of 5’
to 3’.
2) Anti-sense
strand
is the DNA template strand that is used by
RNA pol to form mRNA. And it is in direction of 3’
to 5’.
3) As we know that for each process there must
be a start point for initiation process, so the bases to left of the start
point is upstream and taking negative
sign while to the right is downstream
and taking positive sign.
Now: steps
of mRNA synthesis include initiation, elongation and
termination.
First:
initiation:
In prokaryotes:
Transcription begins with the binding of the RNA pol holoenzyme (sigma
factor) to a region of the DNA known as the promoter, which is not transcribed.
There are
some consensus sequences within promoter region that are
recognized by RNA polymerase holoenzyme:
a) -35
sequence:
A consensus sequence (5-TTGACA-3), centered
about 35 bases to the left of the
transcription start site, is the initial point of contact for the holoenzyme.
b) Pribnow
box: A second consensus sequence (5ʹ-TATAAT-3ʹ),
centered at about 10, which is the
site of initial DNA melting (unwinding).
In eukaryotes:
promoter has 2 characteristic sequences:
a) CAAT
sequence
(-70 sequence) which is centered about 70 bases to the left of the transcription
start site.
b) Hongess
box or TATA
(-25 or -30 sequence) which is centered about
25 or 30 bases to the left of the
transcription start site.
Second:
elongation:
Once the promoter
region has been recognized and bound by the holoenzyme, local unwinding of the
DNA helix continues, mediated by the polymerase.
RNA pol
begins to synthesize a transcript of the DNA sequence, and several short pieces
of RNA are made.
RNA pol uses
nucleoside triphosphates as substrates and releases pyrophosphate each time a
nucleoside monophosphate is added to the growing chain.
Third:
Termination:
The
elongation of the single-stranded RNA chain continues until a termination
signal is reached. Termination can be intrinsic (spontaneous) or dependent upon
the participation of a protein known as the ρ (rho) factor.
a) ρ
-independent termination: Here sequences of mRNA are self-complementary.
This allows the RNA to fold back on itself, forming a GC-rich stem (stabilized
by hydrogen bonds) plus a loop. This structure is known as a “hairpin. And once hairpin is formed, RNA
polymerase stops the process.
b) r
-dependent termination: This requires the participation of an additional protein, rho (r)
where rho binds a C-rich “rho recognition site” near the 5ʹ-end of the nascent
RNA and moves along the RNA until it reaches the RNA pol paused at the
termination site.
Notes:
1) Some antibiotics prevent bacterial cell
growth by inhibiting RNA synthesis. For example, rifampin
(rifampicin) inhibits transcription
by binding to the β subunit of prokaryotic RNA
pol, and preventing chain extension (elongation).
2) Rifampin is used in treatment of TB and box virus.
3) Dactinomycin binds to the DNA
template and interferes with the movement of RNA pol along the DNA.
After we
talk about transcription in prokaryotes, we will illustrate some points of this
process in eukaryotes.
Nuclear
RNA polymerases of eukaryotic cells:
a) RNA
polymerase I: This
enzyme synthesizes the precursor of the rRNA in the nucleolus.
b) RNA
polymerase II: This
enzyme synthesizes the nuclear precursors of mRNA and small ncRNAs
(non-coding mRNA), such as snRNA (regulation of gene).
c) RNA
polymerase III: This
enzyme synthesizes tRNA, 5S rRNA (small subunit), and some snRNA.
Transcription
process in eukaryotes:
Eukaryotic
transcription involves separate polymerases for the synthesis of rRNA, tRNA,
and mRNA. In addition, a large number of proteins called transcription
factors (TFs) which bind to distinct sites on the DNA either within the
core promoter region, close (proximal) to it, or some distance away (distal).
They are required both for the assembly of a transcription complex at the
promoter and the determination of which genes are to be transcribed.
Role
of enhancers in gene regulation:
Are special
DNA sequences that increase the rate of initiation of transcription by RNA pol
II.
They can be
located upstream (to the 5ʹ-side) or downstream (to the 3ʹ-side)
of the transcription start site, and be close to or thousands of base pairs
away from the promoter.
Inhibitors
of RNA polymerase II:
α-Amanitin, a potent toxin produced by the poisonous
mushroom Amanita phalloides (sometimes called “the
death cap”), forms a tight complex with RNA
pol II, thereby inhibiting mRNA synthesis.
Wait for us in another topic in Biochemistry!!!!!!!!
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