PROTEINS AND TRANSLATION.pptfIIIt

TRANSLATION AND PROTEIN SYNTHESIS : 

TRANSLATION AND PROTEIN SYNTHESIS Flordeliz R. Estira MMSU

WHAT IS TRANSLATION? : 

WHAT IS TRANSLATION? Translation is also called protein synthesis which is the RNA directed synthesis of polypeptides.

CENTRAL DOGMA OF MOLECULAR BIOLOGY : 

CENTRAL DOGMA OF MOLECULAR BIOLOGY DNAREPLICATION: DUPLICATION Mechanism: Complementary base pairing Binding and inhibition of RNA polymerase to promoter site DNA TRANSCRIPTION: COPYING Mechanism: a strand of DNA is copied into a new strand of RNA mRNA TRANSLATION: CODING Process by which the information (genetic message) in an RNA (template) is decoded on the ribosomes into specific amino acid sequence in a polypeptide chain ( PROTEIN)

Role of proteins : 

Role of proteins Proteins are used to build cell structures and used as enzymes Through the action of proteins ( directly or indirectly ), the information stored in the DNA (gene ) is expressed ( phenotype, trait )

Slide 7: 

Protein synthesis or translation requires the three types of RNA: 1. Transfer RNA (tRNA) 2. Messenger RNA (m RNA) 3. Ribosomal RNA (rRNA)

Function of each type of RNA : 

Messenger RNA (mRNA) is the blueprint or template for construction of a protein Ribosomal RNA (rRNA) is the construction site where the protein is made Transfer RNA (tRNA) is the vehicle delivering the proper amino acid to the site at the right time. Function of each type of RNA

TRANSLATION : 

During translation, gene sequences in the messenger RNA (mRNA) are used to manufacture proteins Translation occurs at special structures in the cell called ribosomes. Ribosomes are the "factories" where RNA is used to manufacture proteins. TRANSLATION

Structure of tRNAs : 

Structure of tRNAs 1. exhibit a cloverleaf-like secondary structure. 2. have a 5'-terminal phosphate. 3. have a 7 bp stem that includes the 5'-terminal nucleotide and may contain non-Watson-Crick base pairs, e.g. GU. This portion of the tRNA is called the acceptor since the amino acid is carried by the tRNA while attached to the 3'-terminal OH group. 4. have a D loop and a TjC loop. 5. have an anti-codon loop. 6. terminate at the 3'-end with the sequence 5'-CCA-3'. 7. contain 13 invariant positions and 8 semi-variant positions. 8. contain numerous modified nucleotide bases.

A ribosome : 

A ribosome is an organelle composed of ribosomal RNA and ribosomal proteins (known as a Ribonucleoprotein or RNP) It translates Messenger RNA (mRNA) into a polypeptide chain (e.g., a protein) It can be thought of as a factory that builds a protein from a set of genetic instructions can float freely in the cytoplasm (the internal fluid of the cell) or bind to the endoplasmic reticulum, or to the nuclear envelope ribosomes are ribozymes, it is thought that they might be remnants of the RNA world

BASIC STEPS IN THE TRANSLATION PROCESS : 

BASIC STEPS IN THE TRANSLATION PROCESS 1. Activation of Amino Acid 2. Initiation of Translation ( Codon Recognition ) 3. Formation and Elongation of Polypeptide (Translocation ) 4. Termination of Translation

Amino acid Activation : 

Amino acid Activation 1st step : ATP + amino acid -> Aminoacyl- AMP + PPi 2nd step: amino-AMP-AMP + tRNA -> amino-tRNA + AMP

Eukaryotic Initiation Factors and their Functions : 

Eukaryotic Initiation Factors and their Functions Initiation FactorActivityeIF-1repositioning of met-tRNA to facilitate mRNA bindingeIF-2ternary complex formationeIF-2AAUG-dependent met-tRNAmeti binding to 40S ribosomeeIF-2B (also called GEF)guanine nucleotide exchange factorGTP/GDP exchange during eIF-2 recyclingeIF-3composed of ~10 subunitsribosome subunit antiassociation, binding to 40S subunitInitiation factor complexoften referred to as eIF-4Fcomposed of 3 primary subunits: eIF-4E, eIF-4A, eIF-4G and at least 2 additional factors: PABP, Mnk1 (or Mnk2)mRNA binding to 40S subunit, ATPase-dependent RNA helicase activity, interaction between polyA tail and cap structurePABP:polyA-binding proteinbinds to the polyA tail of mRNAs and provides a link to eIF-4GMnk1 and Mnk2eIF-4E kinasesphosphorylate eIF-4E increasing association with cap structureeIF-4AATPase-dependent RNA helicaseeIF-4E5' cap recognition4E-BP (also called PHAS)3 known formswhen de-phosphorylated 4E-BP binds eIF-4E and represses its' activity, phosphorylation of 4E-BP occurs in response many growth stimuli leading to release of eIF-4E and increased translational initiationeIF-4Gacts as a scaffold for the assembly of eIF-4E and -4A in the eIF-4F complex, interaction with PABP allows 5'-end and 3'-ends of mRNAs to interacteIF-4Bstimulates helicase, binds simultaneously with eIF-4FeIF-5release of eIF-2 and eIF-3, ribosome-dependent GTPaseeIF-6ribosome subunit antiassociationback to the top

Specific Steps in Translational Initiation : 

Specific Steps in Translational Initiation 1. A ribosome must dissociate into its' 40S and 60S subunits. 2. A ternary complex termed the preinitiation complex is formed consisting of the initiator, GTP, eIF-2 and the 40S subunit. 3. The mRNA is bound to the preinitiation complex. 4. The 60S subunit associates with the preinitiation complex to form the 80S initiation complex.

Slide 19: 

Several Antibiotic and Toxin inhibitors of Translation

The WOBBLE Hypothesis : 

The WOBBLE Hypothesis

In the process of translation…… : 

In the process of translation…… There are 20 essential amino acids, that can be combined in any order, just like the four nucleotides This permits the production of the many different proteins which let organisms grow and function.

Slide 22: 

Each of the 20 amino acids from which the protein is to be built must be attached to its specific transfer RNA (tRNA) molecule Actually, only the tRNA molecule, and not its attached amino acid, determines where the amino acid is added during protein synthesis Specific enzymes called aminoacyl-tRNA synthetases couple each amino acid to its appropriate tRNA molecule.

THE GENETIC CODE : 

THE GENETIC CODE

Order of Events in Translation : 

Order of Events in Translation 1. synthesis proceeds from the N-terminus to the C-terminus of the protein. 2. the ribosomes "read" the mRNA in the 5' to 3' direction. 3. active translation occurs on polyribosomes (also termed polysomes). This means that more than one ribosome can be bound to and translate a given mRNA at any one time. 4. chain elongation occurs by sequential addition of amino acids to the C-terminal end of the ribosome bound polypeptide.

TRANSLATION : 

The ribosome binds to the mRNA at the start codon (AUG) that is recognized only by the initiator tRNA During this stage, initiation complexes, composed of an amino acid linked to tRNA, sequentially bind to the appropriate codon in mRNA by forming complementary base pairs with the tRNA anticodon The ribosome proceeds to the elongation phase of protein synthesis The ribosome moves from codon to codon along the mRNA Amino acids are added one by one, translated into polypeptidic sequences dictated by DNA and represented by mRNA At the end, a release factor binds to the stop codon, terminating translation and releasing the complete polypeptide from the ribosome. TRANSLATION

Slide 28: 

The Shine-Delgarno Element The Shine-Delgarno element is found at the 5' side of each initiator AUG codon in prokaryotic polycistronic mRNAs. This element is complementary to sequences present near the 3'-end of the 16S rRNA of the prokaryotic ribosome.

Elongation : 

Elongation The elongation phase of protein synthesis on a ribosome requires a three-step cycle which is repeated over and over during the synthesis of a protein chain 1. An aminoacyl-tRNA molecule binds to the A-site on the ribosome 2. A new peptide bond is formed as the ribosome moves a distance of three nucleotides along the mRNA chain and tRNA moves to the P site 3. Ejection of an old tRNA molecule and "resetting" the ribosome so that the next aminoacyl-tRNA molecule can bind.

TRANSLATION : 

TRANSLATION

TRANSLATION : 

TRANSLATION

Slide 41: 

The binding of many ribosomes to an individual mRNA molecule generates polyribosomes

Heme Control of Translation : 

Heme Control of Translation

At a resolution of 30 angstroms, this 3-D model of the eIF3 protein complex shows it be a particle consisting of five lobes - analogous to a head, and a pair of arms and legs. : 

At a resolution of 30 angstroms, this 3-D model of the eIF3 protein complex shows it be a particle consisting of five lobes - analogous to a head, and a pair of arms and legs.