Download To begin with, all the DNA polymerases either the five types in

Genetics
Sheet 3
To begin with, all the DNA polymerases either the five types in
prokaryotes or the nine types in eukaryotes have the 5' to 3' polymerase
activity. Some have other functions. For example: δ polymerase 3'-5' and may b
5'-3' exonuclease activity.
N.B: exo or endonuclease it doesn’t really matter bcz nuclease means
breaking down, exo or endo depends on the side of attack.
δ polymerase is the main one in eukaryotes which has other functions:
- helicase : opens the double stranded DNA then starts copying. This
feature is not found in polymerase III in prokaryotes.
- δ polymerase and polymerase III have high processivity which is the
velocity and capability of working " sor3etoh wi nafasoh fi alsho3'l."
This
means how many nucleotides are added and then it gets tired.
- High fidelity "accuracy" meaning that how many errors it makes which
is one mismatch for 10 billions to 100 billions.
As we know, our DNA is 6 billion base. There is no one polymerase copies all
this 6 billion. We have many bubbles and every polymerase works in a bubble.
The area between a bubble and another is nearly 10000 or 100000 base. A
polymerase may be given a task of 100000 segment at most which is nothing
with its fidelity. It is 0 %.
While if we talk about pol. III, the fidelity is one per 10000. So, 3000
genes >> 3 million nucleotides >> the rate of having errors during copying is
high (may be tens or hundreds, some may be fixed, others may not.)
Other polymerase enzymes: plz read them from the slides
Some notes:
* α polymerase has primase activity → makes the primer for δ
polymerase.
In the previous figure, we can see the primase domain made by α pol.
Which is followed by δ pol.
So, when the primer is being made, primase comes along with α pol.
Primase makes the primer. α pol. is not only bringing the primase, it also
makes 10 – 20 segments as being a polymerase. After that, the δ pol.
comes and takes the responsibility either it is the Okazaki fragments or
the leading strand.
For a long time, it was thought that α pol. synthesizes the Okazaki
fragments for the following reasons:
1- They saw it at first with primase but they didn’t know it would go
soon and replaced by δ pol.
2- Okazaki length is almost 200 – 250 nucleotides which equals the
processivity of α pol. So it might do it.
But after that they discovered that δ pol. synthesizes both.
* γ pol. : it is found in mitochondria.
Mitochondria look highly like bacteria. It is thought that mitochondria
originally are bacterial infection coexisted with our cells millions of years
ago. The evidences supporting this theory are:
- Mitochondria are similar in size to bacteria.
- Mitochondria have double membrane like most of the bacteria.
- DNA of mito. is circular like that of prokaryotes.
- DNA of mito. has no junk (non-coding) like that of bacteria.
- DNA has no introns.
- Even the promoter and γ pol. are similar to that of prokaryotes.
Transcription
Before we go, we have to be familiar with the differences between DNA
replication and DNA transcription:
DNA replication
- DNA synthesis from DNA template
- the two strands are used as templates
to synthesize two strands
- DNA pol. is used which has high
fidelity and needs a primer
DNA transcription
- RNA synthesis from DNA template
- Only one strand is used to synthesize
one new RNA strand similar to the
coding strand
- RNA pol. is used which has low fidelity
and doesn’t need a primer
N.B: that the RNA pol. has low fidelity is not a big deal !!
Bcz there will be nearly a million of copies of RNA from the DNA. So, if
you have errors in ten out of a million ….. so what ?!!
TYPES OF RNA :
( plz dear colleagues read them from the slides.)
Here are some additional notes:
* about mRNA:
- splicing: eliminating the introns and joining the exons together.
- capping functions: protects the RNA from degradation and the most
important function is helping the ribosomes to recognize that this is the 5'
end.
The cap is GTP; methylated guanine. When the pol. is working ,
triphosphate comes which will be splitted and the monophosphte is used.
The only nucleotide that has triphosphate is the GTP of the cap.
- Poly A tail: adding 200 – 250 adenyl groups. It has two functions:
1- Some proteins transport the RNA from the nucleus to the cytoplasm.
They recognize it from the poly A tail; the longer the poly A tail, the
better they recognize it.
2- The RNA has short half life in cytoplasm. It will be broken down by
RNAses so proteins won't be made for ever. But at the same time it
must not be too short. Usually, RNAses start chewing the RNA from
the 3' end. Poly A tail will give some time to ribosomes to translate
enough protein. So, poly A tail helps in prolonging the half life of
mRNA.
* about tRNA:
- it is the true translator not the ribosomes.
- it is not straight but folded. For instance, if it starts with AAA and ends
with UUU, they will make hybridization.
tRNA will form stems and loops by its shape which is called hairpin stem
loop ( look at slide 5 plz.) each tRNA has three loops, the important is the
second one bcz it has three exposed nucleotides which form the
anticodon. Each codon will meet anticodon which has a specific amino
acid.
Aminoacyl tRNA synthetases: are enzymes that help in the recognition
between anticodons and suitable amino acids. When tRNA has the amino
acid it is called charged tRNA. The bond between them is an ester bond.
* about rRNA:
- It has structural and functional roles:
Structural: part of the ribosomal subunit.
Functional: one of the rRNA in the large subunit has catalytic activity;
an enzyme called peptidyl transferase. This contradicts the rule that says
all the enzymes are proteins. This enzyme connects the amino acids
together by a peptide bond.
- species specific:
The size, number, and sequence of rRNA which found in all creatures
specific and constant.
In human being:
We have one of them has a 28s , 18s and 5s …
S : it means sedimentation rate ( measurement of the size of rRNA)
Note:
If the size of rRNA is high > the s is high.
And if the size is low > the s is low.
The enzyme which transcribes all types of RNAs from the DNA is RNA
polymerase.
The differences between RNA and DNA polymerases:
1- All of them synthesize nucleotides , but in the RNA pol. There are no
"DNTPs" but NTPs.
2- The RNA pol. doesn't use primer.
3- Fidelity in RNA pol. is a little bit less.
4- RNA pol. makes a transcription of one strand "template strand."
-There are differences bet. RNA pol. of prokaryotes and eukaryotes:
* The prokaryotes have:
Five subunits " 4 of them called CORE RNA pol. and with the fifth one
are called holoenzyme"
2 alpha, 2 beta and the fifth is sigma.
The 4 subunits alone are enough to make the transcription process.
The role of the sigma subunit is to ride them to the right position " to a
promoter of the gene and if the promoter is ON the gene will be
transcribed and if it is OFF the gene will not be transcribed."
Notes:
- Before the beginning of each gene there is a promoter and before its end
there is a terminator " RNA pol. doesn't know when to start but knows
when to stop."
- There are more than one sigma subunits and each one specialized to
identify a specific gene "we may have 10 genes use the same sigma
subunit."
- In the transcription process:
there is a promoter and terminator
- In the translation process:
there is a start and stop codons.
* In the eukaryotes:
There is no sigma subunit. Instead we have transcription factors. Maybe
we have 30 to 40 trans. factors in one gene.
"although prok. have trans. factors, they are more complex in eukaryotes"
We have more than one RNA pol. One of them is for rRNA the other is
for mRNA which is Pol. 2.
pol. 3 is for tRNA and one kind of rRNA.
NOTES:
- The RNA pol. doesn't transcribe the promoter but it leaves 5 to 10 base
pairs then it starts.
The first one which it transcribes after the promoter we give it No. +1
"transcription start side ", then the second +2 ….
The first one before the trans. start side we give it No. -1 and the second
-2 ……
The importance of numbering is to know the positions.
E.g:
The promoter locates bet. -10 and -100
The direction from the transcription start side to 5' end we call it "up
stream" and from the transcription start side to 3' end we call it "down
stream."
We can also use the terms proximal and distal.
E.g:
+1 is down stream to the promoter and +10 is also down stream to it, but
the +1 is proximal to the promoter and +10 is distal to it.
Before any gene we have 5' regulatory region and the main content is the
promoter which has more than one element "the most important and
consensus element is the TATA box which is found bet. -20 and -30 and
the sequence of it is TATATAA ..
Enhancer:
It is part of the 5' regulatory region and it enhances the function of the
promoter "to promote the trans"
Note:
If the promoter works alone, it will make 100 copies, but if it works with
the enhancer "by trans. factors" it will make 1,000,000 copies by
recruiting more RNA pol. to transcript more.
The promoter has consensus sequence and consensus position but the
enhancer doesn't.
The termination signal of the gene in the eukaryotes called poly A signal
which has the sequence AATAAA.
Actually, the RNA pol. doesn't stop exactly in AATAAA but it
transcribes 30 to 40 gene after poly A signal then stops.
After the transcription of 30 to 40 gene, an enzyme comes to cut at 20.
After that an enzyme called "poly A polymerase" puts poly A tail in the
3'.
There are primary RNA (or hnRNA " heteronuclear RNA" because they
are mixed introns and exons and it is still in the nucleus)
and mature RNA.
The hnRNA will undergo 3 processing events to be converted to mature
RNA that will exit to the cytoplasm and then be translated
The 3 processing events are:
1- capping
2- poly adenylation
3- splicing
Note that the first process is capping.
The end …
A7la ta7ieh le: 3abood Abu Dhaim, M7mad Ra9e3, Raed Alba3dani
be3'iabehm ;)
We are so sorry 4 delaying the sheet but it was really " 5arej 3an
eradatna"
Special thanx to our great doctor: Said Ismail
Dedicated to u all 
Done By:
Mokhtar Alwarafi
Belal Numan