Translation

z Process of converting information stored in

nucleic acid sequences into proteins
z Sequences of mRNA (messenger RNA) are
Biogenesis translated into unique sequence of amino acids
in a polypeptide chain (linear order is preserved
Translation
throughout !)

Translation What is necessary?

z Takes place in the cytoplasm z Template - mRNA

z Exception are few proteins coded by mitochondrial z tRNAs (transfer RNAs)
and chloroplastic DNA z Linked to amino acids
z Performed on ribosomes z Ribosomes
z Many accessory proteins
z Some energy (GTP hydrolysis)

mRNA Genetic code

z Single stranded molecule of RNA that encodes z Codon – specifies the sequence of amino acids
sequence of the polypeptide z Initiation (start) codon
z Transcribed and processed in the nucleus and z AUG – methionine
then exported into cytoplasm z Every protein in a cell starts with methionine
z 5’ end has binding sites for translation initiation z Termination (stop) codons
z Middle is a coding sequence z UAA, UGA, UAG
z 3’ end regulates stability of mRNA

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Genetic code Genetic code

z Universal
z Degenerate - some amino acids are specified by
more than one codon
z 64 possible codons and only 20 amino acids

Eukaryotic mRNA tRNA

z Codes for one protein z Deliver amino acids to the

z Is capped on both 5’ and 3’ translational complex
ends z Serve as adapters between
z 5’ cap - methylated base codons in mRNA and amino
z Protects from nucleases acid
z 4 stems and 3 loops
• 3’ poly-A tail z Anticodon – decoding triplet -
• 50-200 adenines added post-transcriptionally localized on the anticodon stem
• Protects mRNA from degradation

tRNA Charging of tRNA

z L-shaped secondary z Linking amino acids to correct t-RNAs

structure z Catalyzed by aminoacyl-tRNA synthetase (aa-tRNA)
z Anticodon and amino acid aa
z Couples an amino acid to its cognate tRNA
are at the opposite arm of z Fidelity of coupling – 20 different synthetases
the L z Two steps
z Activation of amino acid
z Transfer of amino acid to tRNA
anticodon

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Charging of tRNA Charging of tRNA

z Activation of amino acid by z Transfer of amino acid from

ATP ⇒ amino acyl AMP amino acyl AMP to 3’ end of
z High energy intermediate tRNA (adenine)
z Covalent bond
z High energy bond prepares
amino acid for the formation of
peptide bond with nascent
peptide

Ribosomes Ribosomes

z RNA-protein complexes (ribonucleoproteins) z Composed of small and large subunit

z Place of translation (protein synthesis) z Subunits bind together for translation
z Abundant in cells that synthesize large amounts z Ribosomes self-assemble without additional
of protein factors
z Structurally and functionally similar among
species (differ between prokaryotes and
eukaryotes)

Ribosomes Ribosomes

z Decoding and synthesis takes z Contain rRNA molecules and

place in the cavity between proteins
subunits z In prokaryotes
z Ribosomes move along mRNA z Large subunit contains rRNA
chain during translation (5S and 23S) and 32 proteins
z New peptide exits through the z Small subunit contains 16S
rRNA and 21 proteins
tunnel in the large subunit

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Ribosomes Ribosomes

z In eukaryotes z Cytosolic (free)

z Large subunit contains rRNA z Bound to ER
(5S, 5.8S and 28S) and 49
proteins
z Also located in mitochondria and chloroplasts of
z Small subunit contains 18S
eukaryotic cells
rRNA and 33 proteins

Free ribosomes Bound ribosomes

z Found in the cytosol z Bound to the exterior of the rough endoplasmic

reticulum
z May exist as a single ribosome or in groups known
as polysomes z Occur in greater number than free ribosomes in
cells that secrete their manufactured proteins
z Occur in greater number than bound ribosomes in
(e.g., pancreatic cells)
cells that retain most of their manufactured protein
in the cytosol

Protein synthesis Initiation

z Initiation z Attachment of initiator tRNA

z Elongation (Met-tRNA) to start codon on
z Termination mRNA and assembly of
ribosomal subunits

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Initiation Initiation

z Several initiation factors assist the process z Cap recognition

z Strongly regulated process z mRNA initiation factors
z Uses energy form ATP or GTP hydrolysis z Helicase (ATPase) unwinds mRNA

Initiation Initiation

z Formation of preinitiation z Unwinding and scanning

complex z Complex scans and binds to start codon
z Initiator tRNA (Met-tRNA) z Dissociation of initiation factors
binds to small ribosomal
subunit
z Initiation factors use GTP
hydrolysis

Initiation Elongation

z Association with large ribosomal subunit z Repetitive cycles of codon directed addition of
aa-tRNA
z Aa-tRNA binding
z Proofreading
z Peptidyl transfer
z Translocation

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Elongation Elongation

z Aa-tRNA binding z Proofreading

z Elongation factor eEF-1 z Aa-tRNA is checked against codon
delivers amino acid-tRNA to z GTP hydrolysis and dissociation of eEF-1 from
ribosome ribosome
z eEF-1 is bound to GTP z Correct amino acid-tRNA retained based on codon-
anticodon pairing
z Incorrect amino acid-tRNA escape from the ribosome

Elongation Elongation

z Peptidyl transfer z Translocation

z Large rRNA catalyzes formation of peptide bond z Binding EF-2-GTP
z Precise orientation and stabilization of the z Movement of peptidyl-tRNA to P
transition state site
z Sliding of mRNA by 3 bases
z tRNA moved to the exit
z GTP hydrolysis and release of
EF-2

Termination Polysomes

z Protein factor eTF binds to stop codon and
catalyzes hydrolysis of last amino acid-tRNA
z Peptide is released from the ribosome
z Ribosomal subunits dissociate
z In eukaryotes the same molecule of mRNA can
be simultaneously translated several times
z Each emerging peptide is synthesized on a
separate ribosome
z Many ribosomes on the same “string” of mRNA
are called polysomes

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 What happens after translation? What happens after translation?

z A newly synthesized polypeptide chain must

z Posttranslational modifications start as soon as
undergo post-translational processing to generate
the nascent polypeptide emerges from the
the final protein
tunnel in large ribosomal subunit
z Posttranslational modifications include z That means these changes happen during translation
z Targeting to the appropriate cell compartment when the peptide has NOT been finished yet and is
z Folding still attached to the ribosome (cotranslationally)
z Addition of sugar chains
z Formation of disulfide bonds