Molecular Biology 2

The genetic information of the cell is stored and transmitted in the nucleotide sequences of DNA. Expression of this genetic information involves two stages:- The first stage is transcription to form mRNA that carries specific and precise messages in the form of codons from DNA to the cytoplasmic sites of protein synthesis.

- The second stage is translation of the nucleotide sequence of a mRNA (Codons) into an amino acid sequence of a protein.

Components of translational process

m RNA as a carrier of genetic information. t RNA as an adapter molecule, which recognizes an amino acid on one end and its corresponding codon on the other end. Ribosomes as the molecular machine coordinating the interaction between mRNA, tRNA, the enzymes and the protein factors required for protein synthesis.

Genetic Code: It is the sequence of nucleotides along the DNA that can be translated into the amino acids of proteins. It is the relation between sequence of nucleotides in DNA (or in m RNA) and the sequence of amino acids in a polypeptide chain.

The genetic code is made up of a collection of codons. Codons are the individual words in the genetic code dictionary. Each codon is composed of 3 nucleotide bases. Codon can be defined as: the sequence of three nucleotide bases on mRNA which determines the type and the position of the amino acid that will enter in the structure of protein molecule.

Characters of genetic code

1- Genetic code is degenerate i.e. multiple codons can code for the same amino acid except tryptophan and methionine. Wobble theory: The 3rd nucleotide in a codon is less important than the other two in determining the specific amino acid to be incorporated 2- Genetic code is unambiguous i.e. each codon specifies no more than one amino acid.

3- Genetic code is non-overlapping and Commaless 4- Genetic code is universal i.e. the same code words are used in all organisms (proand eukaryotes) with exception in mitochondria

Protein Biosynthesis
Phases: 1- Initiation 2- Elongation 3- Termination

Initiation of protein biosynthesis

For initiation of protein biosynthesis, there must be:- tRNA - rRNA - mRNA - Eukaryotic initiation factors (eIFs). - GTP, ATP and different amino acids.

t RNA charging:It means recognition and attachment of the specific amino acid to the 3` hydroxyl adenosine terminus ( to the sugar) of tRNA in an ester linkage.

Initiation involves 4 steps:1. Ribosomal dissociation 2. Formation of pre initiation complex 3. Formation of 48 S initiation complex 4. Formation of 80 S initiation complex

II. Elongation
It is a cyclic process involving 3 steps
1.

Binding of aminoacyl tRNA to the A site

2. Peptide bond formation 3- Translocation

Termination

Protein maturation
I- Protein folding II- Post translational processing: 1- Proteolysis 2- Modifications of individual amino acids 3- Addition of certain groups:

Inhibitors of protein biosynthesis

Streptomycin inhibits initiation of prokaryotic protein biosynthesis. Tetracyclines block the A site on prokaryotic ribosome, thus inhibit the binding of amino acyl t RNA. Chloramphenicol blocks peptidyl transfer on prokaryotic ribosomes. Erythromycin inhibits the translocation reaction of prokaryotic protein synthesis.

Other antibiotics are not clinically useful because they inhibit protein synthesis in both prokaryotes as eukaryotes (as puromycin) or only in eukaryotes (as cycloheximide). Puromycin is a tyrosinyl tRNA analog. It can bind to ribosomal A site and prematurely terminate polypeptide synthesis. Cycloheximide inhibits peptidyl transferase in 60 S eukaryotic ribosomal subunit.

Regulation of Gene Expression

Types of Gene Expression: 1- Constitutive gene expression: It is unvarying expression of a gene. It is responsible for expression of House Keeping genes.

2- Regulated gene expression: - It is the expression of genes which the cellular level of their products varies in response to molecular signals. -They are called inducible genes or Repressible genes.

Inducible genes: They are the genes which their products increase in concentration under particular molecular circumstances, i.e.; positive regulation. Repressible genes: They are the genes which their products decrease in concentration in response to molecular signals.

Regulation of gene Expression in Eukaryotic cells:

I- Alteration in genes: 1- Gene loss. 2- Gene Amplification. 3- Gene Rearrangement

II- Transcriptional Regulation: 1- Chromatin Remodelling: a- Methylation b- Histone acetylation and deacetylation

Regulated Expression elements

Basal expression elements

+1 start site

Other regulatory elements

Enhancer and silencer elements

Upstream element (CAAT box or GC box

Proximal element (TATA box)

Coding region

3- DNA regulatory factors: 1- Zing finger motif 2- leucine zipper motif 3- helix-turn-helix motif

III- Post-Transcriptional Regulation:

1- Alternative splicing and polyadenylation sites 2-Regulation of RNA stability 3-RNA editing

IV- Regulation at the level of Translation V- Post-Translational Regulation

Gene Mutation
Definition: Mutations are permenant changes in a DNA sequence. This altered DNA sequence can be reflected by changes in the base sequence of mRNA, and, sometimes, by changes in the amino acid sequence of a protein. Mutations can cause genetic diseases

Types of mutation
I- Point mutation (single base substitution): 1- Transition: In which one purine is 1- Transition: In which one purine is replaced by replaced by another purine or one another purine or one pyrimidine is replaced by another pyrimidine. pyrimidine is replaced by another

pyrimidine. or C (e.g. A G

(e.g. A G or C T)

T)

2- Transversion: in which a purine is replaced by a pyrimidine or a pyrimidine is replaced by a purine. (e.g. T A T C A) G, C G

Effects of point mutation

1- silent mutation: The codon containing the changed base may code for the same amino acid. For example, if the serine codon UCA is given a different third base (to become, say, UCU), it still codes for serine. Therefore, this is termed a silent mutation without any effect on protein structure.

2. Missense mutation: The codon containing the changed base may code for a different amino acid. The substitution of an incorrect amino acid may result in three variable effects on protein structure (e.g. hemoglobin -chain).

a- Acceptable missense mutation AAA Lysine AAU (codons) Asparagine (amino acid) 61

(apparently normal functional hemoglobin). b- Partially acceptable missense mutation GAA GUA Glutamic acid (codons) Valine (amino acid) 6

(Hb S; it can bind and release O2 although abnormal).

C- Unacceptable missense mutation: CAU UAU (codons) Histidine Tyrosine (amino acid) 58 (Hb M; it cannot transport O2).

II. frame shift mutation: It results from deletion (removal) or insertion (addition) of one or more nucleotides in DNA that generates altered m RNAS with different effects on protein structure.

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