Translation Flashcards
*Creation of protein molecules using mRNA as the template
*To translate the nucleotide sequence of mRNA into the amino acid sequence of protein
Translation
How many substrates are in protein synthesis
20 amino acids
- Enzymes and protein factors:
initiation factor (IF), elongation factor (EF), releasing factors
- is the template for the protein synthesis
mRNA
Prokaryotic mRNA is ______; that is, a single mRNA molecule may code for more than one peptides
polycistron
- Eukaryotic mRNA is ______; that is, each mRNA codes for only one peptide.
monocistron
- Is a sequence of three adjacent nucleotides on mRNA that corresponds with a specific amino acid
Genetic codon
Stop codons
UAA, UAG, UGA
Start codon (methionine)
AUG
- 61 codons for 20 amino acids
- A portion of a DNA sequence that does not include a stop codon
Open reading frame
Properties of genetic code:
- Commaless = genetic codons should be read continuously without spacing or overlapping.
- Degeneracy = More than one codon can specify the same amino acid (degenerated codons differ on the third nucleotide).
- Universal = genetic codons for amino acids is the same in humans, animals, plants and bacteria
Refers to the insertion or deletion of nucleotide bases in numbers that are not multiples of three
Frameshift mutation
Non-Watson-Crick base pairing is permissible between the 3rd nucleotide of the codon on mRNA and the first nucleotide of the anti-codon on tRNA
Wobble
in tRNA, which has the proofreading ability
Aminoacyl-tRNA synthetase
- Prokaryotic Met-tRNA^met can be formylated to fMet-tRNAi^met.
For prokaryotes:
* fMet-tRNAi^met can only be recognized by initiation codon.
*Met-tRNAe^met is used for elongation.
For eukaryotes:
*Met-tRNAi^met is used for initiation.
*Met-tRNAe^met is used for elongation.
*Is the place where protein synthesis takes place
Ribosome
Three sites on ribosomes
- Aminoacyl site (A site) = Composed by large and small subunit; Accepting an aminoacyl-tRNA.
- Peptidyl site (P site) = Composed by large and small subunit; Holds the growing amino acid chain.
- Exit site (E site) = Only on large subunit; Releasing the deacylated tRNA
The direction of the protein synthesized:
N-terminal—C-terminal
Protein synthesis process steps
–Initiation
–Elongation
–Termination
Three initiation factors in prokaryotes: IF-1, IF-2 and IF-3
ribosomal binding site in bacteria
Shine-Dalgarno (SD sequence)
- The IF-1 and IF-3 bind to the 30S subunit, making separation between 50S and 30S subunit.
- The mRNA then binds to 30S subunit.
*The complex of the GTP-bound IF-2 and the fMet-tRNA enters the P site. - The 50S subunit combines with this complex.
- GTP is hydrolyzed to GDP and Pi.
- All three IFs depart from this complex.
Elongation
*Three steps in each cycle:
–Positioning an aminoacyl-tRNA in the A site—Entrance
–Forming a peptide bond—Peptide bond formation
–Translocating the ribosome to the next codon—Translocation
*Elongation factors _____and _____ in
bacteria are required.
EF-Tu (elongation factor thermos unstable)
EFG (a translocase enzyme)
Peptide bond formation occurs at A side catalyzes by which enzyme
peptidyltransferase
Step 3: Translocation
*GTP-bound EF-G provides the energy to move the ribosome, one codon toward the 3’ end on mRNA.
*After the translocation, the uncharged tRNA is released from the E site.
To terminate the translation, _____ binds to the last tRNA at the P site
Release factor
- Prokaryotes have 3 release factors:
–RF-1: recognizes UAA and UAG
–RF-2: UGA and UAA
–RF-3: hydrolyses and causes the release of ribosomal subunits (30S and 50s subunit).
- Eukaryotes have only 1 releasing factor:
eRF
eukaryotic initiation
- Four steps:
–Separation between 60S and 40S
subunit
–binding Met-tRNAimeton the 40S subunit
–Positioning mRNA on the 40S subunit
–Associating the 60S subunit
Eukaryotic initiation factor:
Facilitates binding of initiating Met-tRNAMet to 40S ribosomal subunit
eIF2
- Consensus recognition sequence (5’-ACCAUGG-3’)
*Often contains the start codon
Kozak sequence
Eukaryotic elongation factors
eEF-1A (Eukaryotic release factor eRF-1 recognizes all three termination codons (UAA, UAG and UGA) and with the help of protein eRF-3, terminates translation )
eEF-Ib
eEF2
Aggregates of numerous ribosomes that are in the process of actively translating mRNA into protein
Polysomes
The macromolecules assisting the formation of protein secondary structure include
–molecular chaperon
–protein disulfide isomerase (PDI)
–peptide prolyl cis-trans isomerase (PPI)
A group of conserved proteins that can recognize the non-native conformation of peptides and promote the correct folding of individual domains and whole peptides.
Chaperons
Modification of primary structure
*Removalof the first N-terminal methionine residue
*Covalent modification of some amino acids (phosphorylation, methylation, acetylation, …)
*Activation of peptides through hydrolysis
Modification of spatial structure
*Assemble of subunits: Hb
*Attachment of prosthetic groups: E.g. glycoprotein
*Connection of hydrophobic aliphatic chains
*The correctly folded proteins need to be transported to special cellular compartments
*Proteins need signal sequence to be transported
Protein targeting
Inhibitor that causes the premature release of nascent polypeptide chains by its addition to the growing chain end.
- It has a similar structure to Tyr-tRNA.
- It works for both prokaryotes and eukaryotes.
Puromycin
*among the most toxic substance known
*The size of few grains of table salt can kill an adult human
*Eliminate a single adenine base from the large ribosomal subunit in all species
Ricin from castor bean oil (Ricinus communis)
Eukaryotic initiation factor:
First factors to bind 40S subunit; facilitate subsequent steps
eIF2B, eIF3
Eukaryotic initiation factor:
RNA helicase activity removes secondary structure in the mRNA to permit binding to 40S subunit; part of the eIF4F complex
eIF4A
Eukaryotic initiation factor:
Binds to mRNA; facilitates scanning of mRNA to locate the first AUG
eIF4B
Eukaryotic initiation factor:
Binds to the 5’ cap of mRNA; part of the eIF4F complex
eIF4E
Eukaryotic initiation factor:
Binds to eIF4E and to poly(A) binding protein (PAB); part of the eIF4F complex
eIF4G
Eukaryotic initiation factor:
Promotes dissociation of several other IFs from 40S subunit as a prelude to association of 60S subunit to form 80S initiation complex
eIF5
Eukaryotic initiation factor:
Facilitates dissociation of inactive 80S ribosome into 40S and 60S subunits
eIF6
*Heat shock protein (HSP)
HSP70, HSP40, and GreE family
*Chaperonin
GroEL and GroES family
Protein synthesis
Unfolded/misfolded proteins
Native proteins
Protein aggregates
Degraded proteins
