TRANSCRIPTIONTRANSCRIPTION

NECESSARY COMPONENTS

•DNA matrix

•DNA-dependent RNA-polymerase

•АТP, GТP, CТP, UТP

•Мg ions

DIFFERENCE FROM REPLICATION

•Only one strand is used as a matrix

•Only the part of DNA is transcribed (not the entire chain)

RNA Polymerase•There are 3 RNA-polymerases in eukaryotes (for mRNA, rRNA, tRNA)

•RNA pol is core of a larger transcription complex

•Complex assembles at one end of a gene (promoter) when transcription is initiated – transcription initiation

•DNA is continuously unwound as RNA pol catalyzes a processive elongation of RNA chain

The Chain Elongation Reaction

•Mechanism almost identical to that for DNA polymerase

•Growing RNA chain is base-paired to DNA template strand

•Incoming ribonucleotide triphosphates (RTPs) form correct H bonds to template

•New phosphodiester bond formed

• Direction 5’-3’

• Speed - 30-85 nucleotides/sec

Initiation and elongationsteps of transcription

The transcription bubble

•RNA poly-merase reaction

•RNA poly-merase reaction

Transcription Termination

•Only certain regions of DNA are transcribed

•Transcription complexes assemble at promoters and disassemble at the 3’ end of genes at specific termination sequences

PROCESSING

•Transcription occurs in the nucleus, translation in the cytoplasm

•Eukaryotic mRNA is processed in the nucleus

•In some mRNA, pieces are removed from the middle and the ends joined (splicing)

•Introns - internal sequences that are removed from the primary RNA transcript

•Exons - sequences that are present in the primary transcript and the mature mRNA

•Specific enzymes cut out introns and join exons - splicing

PROCESSING

Primary transcript

mRNA

transcription

splicing

DNA

exons exones

7-methylguanosine (CAP)

Poly-A (TAIL)

5’ 3’

RNA

introns

PROTEIN PROTEIN SYNTHESISSYNTHESIS

GENETIC CODE - sequence of mononucleotides in mRNA that specifies the sequence of amino acids in peptide chain

CODON – mRNA triplet base sequence responsible for 1 amino acid

PROPERTIES OF GENETIC CODE

1. Unambiguous. In any organism each codon corresponds to only one amino acid.

2. Code is degenerate. There are multiple codons for most amino acids.

3. Universal. Codons are the same for all organism.

4. Without punctuation. There are no punctuations between trinucleotides.

5. Nonoverlapping. Codons do not overlap each other.

Structure of tRNAs

ANTICODON – triplet in tRNA that can complementary bind to codon of mRNA.

Such base pairing between codon and anticodon is responsible for the translation of genetic information from mRNA to protein.

STAGES OF TRANSLATION

• 1. Recognition• 2. Initiation

• 3. Elongation• 4. Termination

R1 CH

NH2

COOH + HO P

O

O

OH

P

O

O

OH

P

O

OH

O Аденозин

R1 CH

NH2

CO P

O

O

OH

Аденозин + H4P2O7O

Aminoacyladenilate

RECOGNITION

Aminoacyl-tRNA-synthetase

Aminoacyladenilate + tRNA aminoacyl-tRNA + AMP

Activation of amino acids

Each amino acid has a specific tRNA

There is specific aminoacyl-tRNA-synthetase for each AA

The structure of tRNA

Initiation of Translation

•The translation complex is assembled at the beginning of the mRNA coding sequence

•Complex consists of: -Ribosomal subunits-mRNA template to be

translated-Initiator tRNA molecule-Protein initiation factors

Initiator tRNA

•First codon translated is usually AUG

•The initiator tRNA recognizes initiation codons

-Bacteria: N-formylmethionyl-tRNA

-Eukaryotes: methionyl-tRNA

Initiation of protein bio-synthesis Methionyl-тRNA binds to P-center

Sites for tRNA binding in ribosomes

There are two centers: peptidyl (P) and aminoacyl (А)

Elongation1) Positioning of the nextaminoacyl-tRNA in the A site 2) Formation of the peptide bound (enzyme – peptidyl transferase) between methionine and AA in A-centre. The residue of methionine is transferred on the amino group of another AA

3) Translocation – shift of ribosome by one codon. Methionyl-tRNA is released from P-centre. Dipeptidyl-tRNA moves from A-centre to P-centre.

Termination of Translation

•Ribosome comes to terminal codon UGA, UAG or UAA

•No tRNA molecules recognize these codons and protein synthesis stalls

•Protein termination factors F-1, RF-2, RF-3 split off synthesized polypeptide from the last tRNA

•Ribosomal complex dissociates

Termina-tion of Trans-lation

POSTTRANSLATIONAL MODIFICATION

1) Preparing of proteins for different functions

2) Direction of proteins to different locations (targeting)

1. Removing of methionine (formylmethionine)

2. Formation of disulfide and other bonds (secondary, tertiary structures)

3. Proteolytic cleavage

4. Modification of amino acid residues:

- Hydroxylation

- Glycosilation

- Phosphorilation

5. Joining of prosthetic groups or cofactors

6. Formation of the quaternary structure

Regulation of the Protein BiosynthesisThe operon model (by Jacob and Monod)

Inhibitors of Transcription

Antibiotics inhibiting protein

synthesis