TRANSCRIPTION
IN PROKARYOTES
GENE EXPRESSION
The genetic information stored in DNA is divided into units
called genes. In order for an organism to function properly and
reproduce, its genes must be expressed at the appropriate time
and place.
The pathway from DNA to RNA and RNA to protein is
conserved in all forms of life and is called the central dogma.
It illustrates two essential concepts: the flow of genetic information from one generation to the
next (replication); and the flow of information within a single cell, a process also called gene
expression
TRANSCRIPTION
Cells cannot make proteins from DNA. They must convert the DNA into RNA in order to make proteins. This process
DNA-directed RNA synthesis is known as transcription.
It is the process of synthesis of RNA by copying the template strand of DNA.
Transcription copies specific genes from DNA to make a complementary strand of RNA. - Only a gene, NOT the
entire DNA strand is transcribed
The enzyme involved in transcription is RNA polymerase. ATP, GTP, CTP, and UTP are used to produce an RNA
complementary to the DNA template. These nucleotides contain ribose rather than deoxyribose
RNA polymerase
n[ATP, GTP, CTP, UTP] ⎯⎯⎯⎯→ RNA + nPPi
DNA template
RNA synthesis proceeds in a 5′ to 3′ direction with new nucleotides being added to the 3′ end of the growing chain at a
rate of about 40 nucleotides per second at 37°C
RNA POLYMERASE
RNA polymerase in prokaryotes are composed of 5 polypeptide subunits
Four of these subunits, denoted α, α, β, and β’, comprise the
polymerase core enzyme. These subunits assemble each time a gene is
transcribed; they disassemble once transcription is complete. The last
subunit called as σ subunit binds to the core enzyme only during
initiation process. The polymerase comprised of all five subunits is called
the holoenzyme.
Each subunit has a unique role:
α-subunits:
the two α-subunits are necessary to assemble the polymerase on the
DNA;
Β and β’-subunits:
the β-subunit binds to the ribonucleoside triphosphate that will become
part of the nascent “recently-born” mRNA molecule; and
the β’ binds the DNA template strand.
σ subunit:
The fifth subunit, σ, is involved only in transcription initiation. It confers
transcriptional specificity such that the polymerase begins to synthesize
mRNA from an appropriate initiation site.
STAGES OF TRANSCRIPTION
INITIATION
ELONGATION
TERMINATION
INITIATION
The holoenzyme binds specifically and tighty to promoter region.
Bacterial promoters have two characteristic features: a sequence of six bases (often TTGACA) about 35 bases
pairs before the transcription starting point and a TATAAT sequence, or Pribnow box, usually about 10 base
pairs upstream of the transcriptional start site also. These regions are called the -35 and -10 sites and are called
consensus sequences.
Once bound to the promoter site, RNA polymerase is able to
unwind the DNA without the aid of helicases. The -10 site is rich
in adenines and thymines, making it easier to break the hydrogen
bonds that keep the DNA double stranded; when the DNA is
unwound at this region, it is called open complex.
A region of unwound
DNA equivalent to about
two turns of the helix
(about 16–20 bases pairs)
becomes the “transcription
bubble,” which moves
with the RNA polymerase
as it proceeds to transcribe
mRNA from the template
DNA strand during
elongation.
Within the transcription bubble, a temporary RNA:DNAhybrid is
formed. As the RNApolymerase progresses in the 3’ to 5’
direction along the DNA template, the sigma factor soon
dissociates from core RNApolymerase and is available to aid
another unit of core enzyme initiate transcription.
ELONGATION
As the RNA polymerase moves along the strand, it adds complementary
nucleotides as dictated by the DNA template, forming the single-stranded
mRNA that reads in the 5 to 3 direction
As elongation of the mRNA continues, single-stranded mRNAis released
and the two strands of DNAbehind the transcription bubble resume their
double helical structure.
The mRNA is made in the 5’ to 3’ direction so it is complementary and antiparallel to the template DNA.
The polymerase continues transcribing until it reaches a termination site and the mRNA transcript is released for
translation. The section of the DNA that has been transcribed is rewound into its original configuration
TERMINATION
Termination of transcription occurs when the core RNA polymerase dissociates from the template DNA.
The end of a gene or group of genes is marked by DNA sequences in the trailer (which is transcribed but
not translated) and the terminator. The sequences within procaryotic terminators often contain
nucleotides that, when transcribed into RNA, form hydrogen bonds within the single-stranded RNA.
This intrastrand base pairing creates a hairpin-shaped loop-and-stem structure. This structure appears to
cause the RNA polymerase to pause or stop transcribing DNA.
There are two kinds of terminators. The first type
causes intrinsic or rho-independent termination.
It features the mRNAhairpin followed by a stretch of
about six uridine residues.
Once the RNA polymerase has paused at the hairpin
loop, the A-U base pairs in the uracil-rich region are
too weak to hold the RNA:DNA duplex together and
the RNA polymerase falls off
The second kind of terminator lacks a poly-U region,
and often the hairpin; it requires the aid of a special
protein, the rho factor (ρ). This terminator causes rho-
dependent termination.
It is thought that rho binds to mRNA and moves along
the molecule until it reaches the RNA polymerase that
has halted at a terminator.
The rho factor, which has hybrid RNA:DNA helicase
activity, then causes the polymerase to dissociate from
the mRNA, probably by unwinding the mRNA-DNA
complex.
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