Download are mRNA

Expression of genetic
information
(Part II)
Definition:
a complex process in which the genetic information in DNA is
decoded and used to specify the manufacture of specific protein in
the cell.
Includes 2 steps:
1) Transcription or Copying
mRNA molecules are synthesized
as complementary copies of DNA
template
2) Translation
Conversion of genetic language
in mRNA molecule (codons) into
amino acid language of protein
Translation
= Protein synthesis:
Copy of genetic
information in
mRNA (codons)
Conversion
into by
Polypeptide
chain (protein)
Requirements
mRNA
Active tRNA
Ribosomes
• What are the kinds of processes that
take place in nucleus and cytoplasm
during gene expression ?
tRNA
(RNA polymerase III)
Nuclear envelope
mRNA (RNA polymerase II)
protein
large
subunit
tRNA
DNA cisterone
mRNA
Nuclear pore
nucleolus
Small
subunit
DNA d.h.
Ribosome:
2 subunits
• Ribosome
= machines of protein
synthesis
Properties:
-Large subunit:
contains 2 binding site.
P= For peptidyl-tRNA (hold
peptide chain).
A= For amino-acyl-tRNA, that
delivers the next amino acid.
-Small subunit
contains mRNA binding site.
- Separate entities are inactive.
- become active when the 2
subunits are linked to mRNA
during translation.
-Polyribosomes (polysomes) are
mRNA -linked clusters of
ribosomal molecules .
Function:
1- translate the information in
mRNA into protein.
2- hold mRNA, amino-acyl tRNA &
polypeptide chain in a correct
orientation during translation.
3- form the peptide bonds between
amino acids
P- site
L. Subunit
A - site
mRNA
S. subunit
mRNA –binding
site
Ribosome
structure
• tRNA
= transfer RNA
3’
5’
• Small molecules that are transcribed
from tRNA gene.
Anticodon
+
Properties:
• Recognized by amino-acyl-tRNA
synthetase & ribosomes.
• Have amino acid acceptor 3’ end.
• Have anticodon that recognizes and links
the complementary mRNA –codon.
Types:
1) initiator tRNA (carries methionine
recognizes & links the start codon; AUG).
2) regular tRNA for other amino acids.
• -there are (40 tRNA, 20 A.A., 61
codons).
Amino acid
acceptor end
Amino acid
+
ATP
ADP + P
Active tRNA
Amino-acyl
– tRNA
synthetase
Structure of tRNA
• Steps of
Protein Synthesis
Anticodon
5’
3’
5’
3’
Methionine
Initiator-tRNA
1) Initiation
- Begins when initiator
complex (initiator tRNA &
small ribosomal subunit)
binds the start codon of
mRNA.
- That followed by formation
of functioning ribosome by
binding of large ribosomal
subunit to initiator
complex
Start codon
5’
3’
Small
subunit
P -site
mRNA
A -site
5’
Small
subunit
mRNA
2) Elongation
1) Begins by occupying A- site by 2nd
tRNA with its amino acid, that
carries anticodon complementary to
the next mRNA –codon.
1st step
5’
Small
subunit
2) Peptide bond formation; Amino acid
is detached from tRNA in P-site &
joined by peptide bond to amino
acid linked to tRNA in A-site.
3) Translocation,
Ribosome moves one codon in
direction 5’ – 3’, that leads to:
- release tRNA from P-site to the
cytoplasm.
- transfer tRNA & the peptide chain
transfer from A- site to P-site.
- A- site become open to receive
another suitable tRNA.
by repeating steps 1, 2, 3 (elongation
cycle), the peptide chain elongates till
the ribosome reaches the stop codon.
2nd step
5’
Small
subunit
3rd step
GTP
GDP+P+E
5’
mRNA
3) Termination
Releasing
factor
Polypeptide
chain
5’
dissociation
At stop codon:
Release factor recognizes and binds the mRNA - stop
codon. It terminates the protein synthesis by releasing:
- Large & small ribosomal subunits
- Polypeptide chain
- tRNA molecule
- Releasing factor
- mRNA
• Posttranscriptional
modification & processing of
mRNA in Eukaryotic Cell
In Eukaryotic cells:
Transcript mRNA is
immature (called premRNA), why?
a) - contains non-coding
regions (introns) between
protein coding regions
(exons).
b) - needs modifications
before it becomes competent
for transport & translation.
• In prokaryotic cell
(bacteria)
- translation is coupled with
transcription,
Protein synthesis begins while the
mRNA is being completed, as
multiple ribosomes attach to the
mRNA to form a polyribosomes
- No modification or processing,
why ?
• Types of modifications
1) Capping:
- Begins when mRNA is about 2030 nucleotides long.(7 –methyl guanosine cap is
added to 5’ end of mRNA).
Importance:
1) Non-capped mRNA can not be
recognized by ribosomes.
2) Protection against degradation.
3) Increase the stability of mRNA.
Half life:
- about 10 hs. In Eukaryotic cell.
- about 2 mins. In prokaryotic cell.
2) Polyadenylated tail
( Poly-A tail)
addition of 100 – 250 adenine
nucleotides to the 3’ end of
completed mRNA.
Importance:
1) Help in passage of mRNA from
nucleus into the cytoplasm.
2) Stabilizes the mRNA against
degradation (increase life span
of mRNA in cytoplasm)
Intron I
3) Splicing:
5’
P-P-P-
= Means removing the
introns (non-coding
regions) and splicing
the exons (coding
regions) together.
1) capping
Intron II
3’-OH
Immature
mRNA
by addition 7methyl Guanosine
to 5” end.
-OH
G-P-P-P-
2) Addition polyadenylate
tail to 3’end.
• Removal of introns is
performed by small
nuclear ribonucleoprotein
complex enzyme
(snRNPC).
G-P-P-P-
-AAAAAA…
3) Splicing the exons
• Function:
formation a continuous
protein coding message.
to form continuous
coding message
Remove
introns I & II
Mature mRNA
G-P-P-P--
--AAAAAA…
Nuclear envelope
Mature mRNA becomes
competent for transport &
translation in cytoplasm
G-P-P-P--
--AAAAAA…
Gene expression in
Eukaryotic cell
1) Transcription & translation
are not coupled.
2) Average half life 10 hours
(translation continuous
about 10 hours).
3) After transcription, mRNA is
modified & processed.
prokaryotic cell
1) Transcription & translation
are coupled.
2) Average half life 2 minutes
3) mRNA is ready for
translation as soon as it
transcribed (no
modification).
Half life = the time that required for deterioration half of
the molecule forming mRNA.
• Characters of genetic code
1) Triplet code (three bases)
2) Universal for all organisms
ex. : UUU = phenylalanine in all
organisms.
3) Redundant, redundancy of
genetic code: more than one
code specify one amino acid. (61
codon, 40 tRNA, 20 amino acid).
4) Reads as a series of nucleotide.
- No comma between the codon
- The start codon determines the
reading frame.
5) The codon could undergo
mutation
The Genetic Code
• Mutations


Definition:
Any change in the nucleotide sequence of DNA.
Two types:
1) Unstable mutation:
Revert back to original sequence.
2) Stable mutation:
Change the characteristics (phenotype) of
organism.
• Mutants
Definition :
An organism showing deviation in some
characters (phenotype), whose progeny
maintain these deviances.
Cause:
molecular change in hereditary material.
Types:
1- Base substitution (Missense & nonsense mutants).
2- Frame –shift mutants.
2) Missense &
Nonsense mutants:
 a)
Missense mutation
Cause: Base substitution mutation in DNA.
Results: one amino acid is replaced with another in a protein.
The effect: depends upon the position of the replaced amino acid.
Missense mutation
(Base substitution
mutation)
The amino acid is not
a part of the active site
No change in
enzyme activity
The amino acid is located
at or near the active site
Reduce or absence the
enzyme activity
( in Sickle cell disease; glutamic
acid is replaced with valine in
Hemoglobin)
b) Non-sense mutation:
Cause: mutation that creates an internal stop codon in a gene,
will prematurely terminates the encoded protein.
5’____ ATG ……………..T A C……………TAG____3’
3’____ TAC …………….. AT G……………ATC____5’
DNA d.h.
mutation
5’____ ATG ……………..T A G……………TAG____3’
3’____ TAC …………….. AT C……………ATC____5’
transcription
5’____ AUG …………….. UAG……………UAG____3’
Start codon
internal stop
codon
mRNA
normal stop codon
translation
Meth. ________________
Protein terminates prematurely
shorter or nonfunctional protein
Base –pair substitution leads to formation internal stop codon (UAG,
UGA, UAA), that replaces the amino acid codon.
3) Frame –shift mutants

Cause:
- Insertion or deletion base – pair.
Results:
- Misreading of all codons from the point of deletion
or insertion to the end of the message.
- Change the amino acid sequence of encoded polypeptide
chain.
AUG ACA CAA UGG ACU GAC ………… mRNA
Meth. ………………………………………… protein (normal)
A
Deletion of a single base
AUG ACA CAU GGA CUG AC ………… mRNA
Meth. ………………………………………… protein (unuseful)
• Results: - formation of unuseful new polypeptide chain.