Master Students

Lecture :2
Assistant. Prof. Dr. Hanaa N. Abdullah

Transcription
-The first step in gene expression transfers the genetic
formation stored in DNA- genes in to mRNA
molecules that carry the information to the
ribosomes- the sites of protein synthesis- in the
cytoplasm.
- The basic features of transcription are the same in
both prokaryotes and eukaryotes, but many of the
details,such as the promoters sequences are
different.

Transcription in Prokaryotes
A-The initiation of transcription
- RNA polymerase bind to ds-DNA at the beginning of the
gene to be copied.
-RNA polymerase recognize and bind to promoters.
-Promoters: specific sequence near the beginning of gene
where the transcription will start.
-In bacteria, RNA polymerase consists of a core enzyme ,
plus a (sigma) subunit involved only in initiation .
- The (sigma) subunit reduce RNA polymerases general
affinity for DNA but simultaneously increases RNA
polymerases affinity of the promoter.

- After binding to the promoters, RNA polymerase

Unwinds part of the double helix, exposing unpaired
bases on the template strand. The complex formed
between the RNA polymerase and an unwound
promoter is called an open promoter complex.
-The enzyme identifies the template stand and chooses
the two nucleotide with which to initiate copying(Base
pairing)
-RNA polymerase aligns the first two ribonucleotides of
new RNA, which will be at the 5 end of gene.
-RNA polymerase that catalyzes the formation of a
phosphodiaster bond between the first two
ribonucleotide.
- Soon thereafter, RNA polymerase releases the(sigma)
subunit. This release marks the end of initiation.

B- Elongation : Constructing an RNA Copy of the gene

-These changes enable the core enzyme to leave the promoter yet
remain bound to the gene.
-The core enzyme now moves along the chromosome , unwinding
the double helix to expose the next single-stranded region of
the template.
-The enzyme extends the RNA by linking a ribonucleotide
positioned by complementary with the template strand to the
3end of the growing chain.
-As the enzyme extends the mRNA in the 5 to 3 direction, it
move in the antiparallel 3 to 5 direction along the DNA
template strand.
-The region of DNA unwound by RNA polymerase is called the
transcription bubble.
- Within the bubble, the nascent RNA chain remains base paired
with the DNA template , forming a DNA-RNA hybrid.

Once an RNA polymerase molecules can move in to initiate

transcription. If the promoter is very strong , that is, if it can
rapidly attract RNA polymerase, the gene can undergo
transcription
by
many
RNA
polymerase
simultaneously.

C- Termination : The End of Transcription

-RNA sequences that signal the end of transcription are known

as Terminators.
-There two types of terminators:*Intrinsic terminators, which cause the core enzyme to
terminate transcription on its own, and
*extrinsic terminator, which require proteins other than RNA
polymerase particularly a polypeptide Known as rho- to
bring about termination.
-All terminators, whether Intrinsic or extrinsic , are specific
sequences in the mRNA that are transcribed from specific
DNA region.
- A terminators often form hairpin loops in which nucleotides
within the mRNA pair with nearby complementary
nucleotide . Upon termination, RNA polymerase and a
completed RNA chain are both released from the DNA.

D- The product of Transcription is single- stranded

primary transcript
-The RNA produced by the action of RNA polymerase

nucleotides known as a on a gene is a single strand of

primary transcript.
-The bases in the primary transcript are
complementary to the bases between the initiation
and termination sites in the template strand of the
gene.
-The riponucleotides in the primary transcript include a
start codon, the codons that specify the remaining
stop codon. amino acids of the polypeptide, and a

Promoters of bacterial genes

Transcription
3
5
Primary transcript

promoters

-35

-10

+1

Transcription in Eukaryotes
- Three different enzyme catalyze transcription in
eukaryotes, and the resulting RNA transcript
undergo 3 important modification , including the
excision of non-coding sequences called introns.
The nucleotide sequence of some RNA transcripts
are modified
posttranscriptional by RNA editing.
- In eukaryotes, RNA synthesized in the nucleus, and
most RNA that encode proteins must be
transported to
the cytoplasm for translation on ribosomes.

- Three types of RNA polymerase are present in

eukaryotes; each enzyme catalyzes the
transcription of specific class of genes.
Enzyme

location

Products

RNA polymerase I

Nucleolus

rRNAs, excluding 5S
rRNA

RNA polymeraseII

Nucleus

Nuclear Pre-mRNA

RNA polymeraseIII

Nucleus

tRNA, 5S rRNA, and other

small nuclear RNAs

1-initiation of RNA chains

Unwound segment of DNA, the formation of the unwound segment of DNA required to initiated transcription factors.
transcription factors: Some 50 different protein(transcription factor bind to
promoter sites, usually on the 5 side of the gene to be transcribed.
-The promoters recognized by RNA polymerase II consist from short
transcription start point. conversed elements located upstream from the
- The conversed elements closest to the transcription start site (+1) is called
consensus TATAAAA (reading 5 to 3on non the TATA box; it has the
template strand) and is centered at about
position -30.
-The TATA box play important role in positioning the transcription start point.
-The second conversed element is called the CAAT box (GGCCAATCT), it
occur near position -80. Two others conversed element , The GC box
(GGGCGG) and the octamer box (ATTTGCAT) often are present in
RNA polymerase II promotors.

- The initiation of transcription by RNA polymerase

II requires the assistance of several basal
transcription factors. The basal transcription
factors must interact with promoters in the correct
sequence to initiate transcription effectively .
- Each basal transcription factors is denoted TFIIX
(transcription factors RNA polymerase II, where X
is a letter identifying the individual factors).
- TFIID is the first basal transcription factors to
interact with promoters .
- Next ,TFIIA joins the complexes.
- Followed by TFIIF and RNA polymerase TFIIB,
TFIIF,TFIIE.

2- RNA chain elongation and termination .

In eukaryotes, the majority of the primary transcripts of
genes that encode polypeptides undergo three
modifications prior to their transport to the cytoplasm
for translation.
1- 7 methyl guanosine caps are added to the 5 ends of
the primary transcripts.)elongation)
2- Poly (A) tails are added to the 3 ends of the
transcripts, which are generated by cleavage rather than
by termination of chain extension.)termination)
3- When present, intron sequences are spliced out of
transcripts.

How RNA processing adds a tail to the 3end of

eukaryotic mRNA.

- The 5 cap on most eukaryotic mRNA is a 7

methyl guanosine residues joined to the initial
nucleotide of the transcript by a 55phosphate linkage.
- The 3 (A)tail is a polyadenosine tract 20-200
nucleotides long.
-

- The mechanism of RNA Splicing .

- The non coding introns are excised from gene transcripts by several mechanisms.
-In the primary transcripts of nuclear genes, the only completely conversed seq. of different introns are dinucleotide sequences at the end s of introns, namely,
Intron
exon-GT..............AG-exon

-The human genome is estimated to contain some 180.000 exon ,

with a current of 21,000 genes, the average of our genes is about 9.
In general, introns tend to be much longer than exons. An average
eukaryotic exon is only 140 nucleotides long, but one human intron
stretches for 480,000 nucleotides.
Removal of the introns- and splicing the exon together are among

the essential steps in synthesizing mRNA.

Genetic code
-Is composed of nucleotide triplets(3 nucleotides) in
mRNA specify one amino acid in the poly peptide
products; thus, each codon contains 3 nucleotides.
-The genetic code contains start codon and stop
codon. Specfic codons are used to initiate and to
terminate polypeptide chains.
-The genetic code is ordered: multiple codons a given
a.a and codons for a.as with similar chemical
properties are closely related, usually differing by a
single nucleotides..

Twenty different a.as are incorporated into polypeptides

different codon during translation. Thus, at least 20
must be formed with the 4 bases available in mRNA.
Two bases/codon would result in only 4= 16 possible
codon- clearly not enough. 3=46 possible codon.
- Intiation and Termination codons.
- In both prokaryote and Eukaryote, the codon (AUGMet) is used to initiate polypeptide chains. In rare
instances,(GUG Valine) is used as an initiation
codon. In eukaryote, the codon must be the first AUGMet encountered by the ribosome as it scans from the
the mRNA molecules. At internal 5 end of
positions, AUG is recognized by tRNA met,and GUG
is recognized by a valine tRNA.

- Three codons (UAG-amber, UAA-ochre and

UGA-opal) specify polypeptide chain
termination.

Types of RNA
mRNA :-This will later be translated into a
polypeptide
rRNA:- This will be used in the building of ribosome:
machinery for synthesizing proteins by translating mRNA.
There are 4 types. In eukaryotes, these are:
-18S rRNA: One of these molecules, along with some 30 different
protein molecules, is used to make the small subunit of the
ribosome.
-28S, 5.8S and 5S rRNA:one each of these molecules, along with some 45 different proteins, are used to make the large subunit of
the ribosome.
- S number given each type of rRNA reflect the rate of the
molecules sediment in the ultracentifuge.
.

Protein Synthesis: Translation

- The genetic information in mRNA molecules is translated into the
amino acid sequences of polypeptides according to the
specifications
of the genetic code.
- The process by which the genetic information in mRNA
molecules is translated into the amino acid sequences of
polypeptides according to the specifications of the genetic code,
into the sequence of amino acids in the polypeptide gene products
is complex , requiring the functions of a large number of
macromolecules. These include:
1-Over 50 polypeptides and three to five RNA molecules present
in each ribosome.
2-At least 20 amino acid-activating enzymes.
3-40-60 different tRNA molecules and
4-numerous soluble proteins involved in polypeptide chain
initiation, elongation, and termination.

The structure of tRNA

-tRNA serve as adaptor molecules that mediate the transfer of

information from nucleic acid to protein.
-short, ssRNA molecules 74-95 nucleotides in length.
-Each tRNA carries one particular amino acid, and cells must have at
least one tRNA for each of the 20 amino acids specified by genetic
code.
-The name of tRNA reflects the amino acid it carries. For example
tRNA GLY carries the amino acid glycine The structure of a tRNA
molecule on 3 level.
1-The nucleotide sequence of a tRNA constitutes the primary
Structure
2- Secondary structure
3-Tertiary structure.

tRNA mediate the transfer of information from nucleic

acid to protein.