Molecular Cell Biology Lecture 4
1. RNA polymerase transcribes DNA into RNA, 5’ to 3’, by binding to specific DNA sequences and
utilizing phosphate bond energy (derived from hydrolysis of the NTP) to drive elongation.
2. Prokaryotic RNA polymerase consists of several subunits and a single type of polymerase conducts
all transcriptional activities.
3. In eukaryotes, three different polymerases are used: Pol I: responsible for synthesis of large
(rRNAs); Pol II: mRNAs; Pol III: small RNAs (e.g. tRNA). Each of these RNA polymerases is a
4. Consensus sequences exist upstream (5’ to) the gene they control and RNA polymerase interacts
and specifically binds to some of these.
5. Most eukaryotic RNAs are synthesized as preRNAs, which are then processed to yield a functional
6. Ribosomal RNA (rRNA) is synthesized at specific sites within the nucleus called the nucleolar
organizer region, which contains many tandemly repeated copies of the rRNA genes.
7. Pulse chase experiments were instrumental in deciphering how processing occurs.
o Transcription is the process in which a DNA strand provides the information for the synthesis of an
The enzymes responsible for transcription in both prokaryotic and eukaryotic cells are called
DNA- dependent RNA polymerases (RNA polymerases)
Enzymes are able to incorporate nucleotides, one at a time, into a strand of RNA whose
sequence is complementary to one of the DNA strands, which serves as the template
The steps in the synthesis of RNA include:
a. First, the association of the polymerase with the DNA template (interaction between two
different macromolecules = proteins + nucleic acids)
The site on the DNA to which an RNA polymerase molecule binds prior to
initiating transcription is called the promotor; contains the information that
determines which of the two DNA strands is transcribed and the site at which
Cellular RNA needs the help of additional proteins called transcription factors
which help recognize the promoters
b. RNA polymerase moves along the template DNA strand toward its 5’ end. As the
polymerase progresses, the DNA is temporarily unwound, and the polymerase
assembles a complementary strand of RNA that grows from its 5’ terminus in a 3’
RNA polymerase catalyzes reaction; RNAn + NTP RNAn+1 + PPi; in which
ribonucleoside triphosphate substrates (NTPs) are cleaved into nucleoside
monophosphates as they are polymerized into a covalent chain
c. Those reactions leading to the synthesis of nucleic acids and proteins must occur under
conditions in which there is virtually no reverse reaction. This condition is met during
transcription with the aid of a second reaction
PPi 2 Pi a pyrophosphate. PPi produced in first reaction is hydrolyzed to
inorganic phosphate Pi the hydrolysis pyrophosphate releases a large amount of
free energy and makes the incorporation of nucleotides essentially irreversible
d. As the polymerase moves along the DNA template, it incorporates complementary
nucleotides into the growing RNA chain.
Nucleotide is incorporated into the RNA strand if it is able to form a proper base
pair with the nucleotide in the DNA strand being transcribed
Consequently, the RNA chain does not remain associated with its template as a
RNA polymerases are apable of incorporating from about 20 to 50 nucleotides
into a growing RNA molecule per secnd, and many genes in a cell are
transcribed simultaneously by a hundred or more polymerases.
e. RNA polymerases are capable of forming prodigiously long RNAs = must remain
attached to the DNA over long stretches = processive
Even though polymerases are relatively powerful motors these enzymes do not
necessarily move in a steady, continuous fashion but may pause at certain
locations along the template or even backtrack before resuming their forward
o Bacteria, such as E.coli, contain a single type of RNA polymerase composed of five subunits that are
tightly associated to form a core enzyme.
If the core enzyme is purified from bacterial cells and added to a solution of bacterial DNA
molecules and ribonucleoside triphosphates, the enzyme binds to the DNA and synthesizes
RNA molecules produced are not the same as those found within cells because the core
enzyme has attached to random sites in the DNA, sites that it would normally have ignored in
If a purified accessory polypeptide called sigma factor is added to the RNA polymerase
before it attaches to DNA, transcription begins at selected locations
The enzymes responsible for transcription in both prokaryotic and eukaryotic cells are called. Enzymes are able to incorporate nucleotides, one at a time, into a strand of rna whose sequence is complementary to one of the dna strands, which serves as the template. The steps in the synthesis of rna include: first, the association of the polymerase with the dna template (interaction between two different macromolecules = proteins + nucleic acids) Cellular rna needs the help of additional proteins called transcription factors which help recognize the promoters: rna polymerase moves along the template dna strand toward its 5" end. As the polymerase progresses, the dna is temporarily unwound, and the polymerase assembles a complementary strand of rna that grows from its 5" terminus in a 3" direction. This condition is met during transcription with the aid of a second reaction.