TRANSCRIPTION

 Transcription is the process by which a DNA template is copied to an RNA strand. The
primary enzyme of transcription is RNA polymerase.
 RNA polymerases are generally composed of several subunits. However, since RNA
polymerase performs essentially the same vital function in all cells, it is not surprising that
RNA polymerases from all organisms share many features.
 Overall, RNA polymerase resembles a crab claw. A channel runs through the enzyme,
starting between the two pincers of the claw. The active site is found at the base of the
pincers within a region called the active centre cleft.
 The active site contains two metal ions. This is consistent with the mechanism for nucleotide
addition described for DNA polymerase. Like DNA polymerase, RNA polymerase always adds
nucleotides to the 3’ end of a growing polynucleotide.
 The transcription cycle is a series of events between the binding of RNA polymerase to the
target gene and dissociation of RNA polymerase and the completed RNA transcript from the
DNA. The transcription cycle is divided into three phases: initiation, elongation and
termination.
 Transcription initiation can be divided into three steps. In the first step, RNA polymerase
binds to a region of the DNA called a promoter. In bacteria, this step involves an initiation
factor called sigma, which recognizes various sequences within promoters. The RNA
polymerase with sigma attached binds the promoter in a defined orientation, so the same
DNA strand is always transcribed from a given promoter. The RNA polymerase and promoter
form a closed complex.
 In the second step, the closed complex undergoes a transition to the open complex. The
pincers at the front of the enzyme clamp down tightly on the downstream DNA. Sigma also
changes conformation, and the DNA strands separate, forming a bubble of single- stranded
DNA.
 In contrast with DNA polymerase, RNA polymerase is able to begin synthesis of a new
polynucleotide without a primer. In bacteria, however, once RNA synthesis begins, the RNA
polymerase goes through a period called abortive initiation. The enzyme synthesizes short
RNA molecules of less than ten nucleotides.
 This is probably because a region of sigma partially blocks the RNA exit channel. Once this
region has been ejected and an RNA chain longer than 10 nucleotides has been synthesized,
the elongation phase begins. Transition to the elongation phase is called promoter escape.
 During elongation, RNA polymerase unwinds the DNA in front of the enzyme, synthesizes
RNA, proofreads RNA, dissociates RNA from DNA, and re- anneals the DNA behind the
enzyme. In contrast with the DNA polymerase, RNA polymerase is able to perform these
functions without the assistance of other proteins.
 The proofreading activity of RNA is less efficient than that of DNA polymerase. While the
proofreading activity of DNA polymerase only allows 1 mistake per 10 million nucleotides,
the proofreading activity of RNA polymerase allows about 1 mistake per 10 thousand
nucleotides.
 RNA polymerase has two proofreading mechanisms. The first mechanism is called
pyrophosphorolytic editing, and involves a simple back- reaction to catalyse the removal of
an incorrectly inserted ribonucleotide. RNA polymerase slows down when an incorrect
ribonucleotide is added, making removal of an incorrect base more likely than removal of a
correct base.
 The second mechanism is called hydrolytic editing, and involves backtracking of the enzyme
by one or more nucleotides to cleave the RNA product. In bacteria, hydrolytic editing is
stimulated by Gre factors, which also act as the elongation stimulating factors.
 Sequences called terminators trigger the elongating polymerase to dissociate from the DNA
and release the completed RNA chain. This animation describes bacterial transcription
termination. Bacteria have two types of terminators: Rho- independent and Rho-
dependent.
 Rho- independent terminators, also called intrinsic terminators, consist of two sequence
elements: a short inverted repeat of about 20 nucleotides, and a stretch of about eight A: T
base pairs.
 The RNA that results from the invert repeat sequence is able to base pair with itself, and
forms a hairpin structure. The hairpin then disrupts the elongation complex. A: U base pairs
are the weakest of all base pairs, allowing the RNA to easily dissociate from DNA.
 Rho- dependent terminators are less well- characterised, but optimally consist of stretches
of about 40 nucleotides that do not fold into a secondary structure in the RNA transcript.
Rho- dependent terminators require the action of the Rho factor, a ring- shaped hexameric
protein.
 The Rho factor binds to the transcribed RNA as it exits the polymerase. Using the energy
derived from ATP hydrolysis, the Rho factor pulls the RNA from the template and RNA
polymerase.
 Since RNA polymerase dissociates the RNA transcript from the DNA as it is transcribed,
multiple RNA polymerases can transcribe the same gene at the same time. This allows a cell
to synthesize a large number of RNA transcripts from a single gene in a short time.

Conclusion

 The primary enzyme of transcription is RNA polymerase. Both prokaryotic and eukaryotic
RNA polymerases resemblea crab claw.
 RNA polymerases are generally composed of several subunits.
 The active site of RNA polymerase contains two metal ions. Just as in DNA polymerisation,
metal ions promote the addition of an incoming nucleotide to 3’ end of the growing RNA
chain.
 The series of events between binding of RNA polymerase to the target gene and dissociation
of RNA polymerase and the completed RNA transcript from the DNA is called the
transcription cycle. The transcription cycle is divided into three phases: initiation, elongation
and termination.
 Transcription initiation can be divided into three steps: binding of RNA polymerase to form a
closed complex, transition to an open complex and promoter escape. In bacteria, an
initiation factor called sigma recognizes various sequences within promoters.
 During elongation, RNA polymerase unwinds the DNA in front of the enzyme, synthesizes
RNA, proofreads RNA, dissociates RNA from the DNA, and re- anneals the DNA behind the
enzyme.
 During termination, sequences called terminators trigger the elongating polymerase to
dissociate from DNA and release the completed RNA chain. Bacteria have two types of
terminators: Rho- independent and Rho- dependent.