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Protein Synthesis
The Building of Proteins from a
Nucleic Acid Template
DNA
• Provides the Template.
• Information contained in the SEQUENCE
of N-bases found along the DNA molecule.
A G C C T A G G G A T A G
T C G G A T C C C T A T C
• Transferring that information into an Amino
Acid SEQUENCE is the trick.
A 2-step Process
1. Transcription
DNA
RNA
2. Translation
RNA
Protein
Transcription
• Production of RNA.
• RNA = Ribonucleic Acid.
– Ribose Sugar.
– Single Stranded Molecule.
– Uracil instead of Thymine.
Transcription
• RNA Polymerase
• Attaches at a Promoter – a sequence of
the DNA that indicates where a gene
starts.
• With the help of Transcription Factors.
Transcription
• A “window” in the DNA is opened –
ultimately, a gene.
• RNA Nucleotides are added (from 5’ to 3’)
in accordance with the DNA parent
template (which is read from 3’ to 5’).
Transcription
• Complimentary
Base Pairing.
• DNA
RNA
G - C
C - G
T - A
A - U
Transcription
• Elongation of a single stranded RNA
molecule.
• The DNA window closes behind.
Transcription
• Termination – a sequence of bases is
reached signaling RNA Polymerase to
release.
• The “gene” closes
• The RNA transcript is released.
RNA Processing
In Eukaryotic cells, things get
a little more complex:
1. Several types of RNA
polymerase are present –
each with a slightly different
function – I, II, and III.
2. A larger mRNA is
transcribed…
RNA Processing
•
•
During Transcription, RNA polymerase II
transcribes MORE than just the proteinencoding part of the gene.
Untranslated Regions (UTRs) are
transcribed at the 5’ and 3’ ends.
AAUAAA
5’ UTR
3’ UTR
– The 3’ UTR is called a Polyadenylation signal.
RNA Processing
• AFTER Transcription is terminated,
additional alterations are made to the ends
of this “pre-mRNA” strand.
5’cap
AAUAAA
Poly-A tail
• A 5’ cap at the leading (5’) end (composed
of a Guanine-like nucleotide).
• A poly-A tail at the trailing (3’) end
(composed of many Adenine nucleotides).
5’ Cap and Poly-A Tail
These END pieces serve several functions:
1. Help “ferry” the eventual mRNA transcript
out of the nucleus.
2. Protect the protein-encoding area from
degradation.
3. Help attach the proper end (5’) of the
transcript to the place where Translation will
take place – the Ribosome.
Further RNA Processing
• RNA splicing – removes portions of the
transcript.
Introns
5’cap
AAUAAA
Poly-A tail
Exons
• Removed segments are called INTRONS.
• The remaining coding segments are called
EXONS.
Further RNA Processing
• INTRONS are
recognized by
snRNPs…
– “small nuclear
RiboNucleoProteins”.
– snRNPs cut out
INTRONS and join the
adjacent EXONS.
– This occurs in the
nucleus.
Further RNA Processing
• The resulting molecule is the RNA transcript
proper (no longer the pre-mRNA).
5’cap
AAUAAA
5’cap
AAUAAA
Poly-A tail
Poly-A tail
Why?
• Introns may be vestigial.
• Introns may have regulatory roles.
• Alternative RNA splicing – more than one
polypeptide from a single gene.
• More chances of unique Exon
combinations (taking into account
“crossing over” during meiosis).
– (Cross-overs at introns won’t disrupt exons).
3 Types of RNA
• mRNA = Messenger RNA
– The “blueprint”
• rRNA = Ribosomal RNA
– The “workbench”
• tRNA = Transfer RNA
– The “truck”
mRNA – messenger RNA
• The “message” – the blueprint for the
production of a polypeptide – a protein.
• But there’s only 4 N-bases, and 20 amino
acids!
• What’s the code?
The Genetic Code
• Triplet Base Code.
• Every 3-letter word in the RNA Transcript
is a CODON.
The Genetic Code
• Each CODON codes for a single Amino
Acid.
The Genetic Code
rRNA – Ribosomal RNA
• The location for protein synthesis – the
workbench on which a polypeptide is built.
• rRNA makes up a RIBOSOME.
• Ribosomes have 2 sub-units.
Large subunit
Small subunit
rRNA – Ribosomal RNA
• The Large subunit has several binding
sites:
tRNA – Transfer RNA
• The “transfer-er” – the
truck that brings
individual Amino Acids
to the workbench for the
production of a
polypeptide – a protein.
• With a driver – the
Anticodon.
tRNA – Transfer RNA
• Each tRNA carries a specific
Amino Acid to the Ribosome.
• It “knows” when to drop this Amino
Acid off by…
• COMPLIMENTARY BASE
PAIRING of its Anticodon with the
corresponding Codon found on
the mRNA.
tRNA – Transfer RNA
• Aminoacyl-tRNA
synthetase = the
enzyme responsible
for hooking Amino
Acids to tRNAs.
• Aminoacyl tRNA =
truck with cargo.
Translation
• Translating the RNA
sequence data into
Protein sequence
data.
• N-bases
Amino Acids
• In the cytoplasm of
Eukaryotic cells.
Translation
• Initiation - All 3 types of RNA come together.
• This requires some energy (provided by
Guanosine Triphosphate -- GTP).
Translation
•
Elongation – Amino Acids are added with
the help of elongation factors (proteins).
1. Codon meets Anticodon
at the ‘A’ binding site.
2. A Peptide Bond Forms
between adjacent Amino Acids
Translation
• Then, a shift in the Ribosome
(a “Translocation”) moves the empty tRNA
to the exit (E-site), opening the A-site for
the next tRNA.
Translation
• Termination…
• Continued elongation until a “STOP”
codon is reached (UAG, UAA, or UGA).
• A release factor (protein) binds to this
codon, effectively releasing the
polypeptide.
• The mRNA, tRNA, and rRNA separate.
Protein Synthesis
Primary Structure
Of the Protein
Protein Synthesis
• In response to a need for a particular
protein – need it NOW.
• The same gene can be transcribed by
several RNA polymerase molecules
simultaneously…allowing faster protein
production.
• Polyribosomes – strings of Ribosomes
translate a single mRNA simultaneously –
allowing faster protein production.
Polyribosomes
Bound and Free Ribosomes
• Free-floating Ribosomes are the sites for
free-floating protein production.
• Ribosomes bound to membranes are the
sites for membrane-bound and secretory
proteins.
Review – Fig. 17.26
Point Mutations
• Point Mutations occur at single nucleotides
along DNA’s template strand.
• Substitutions – the replacement of a pair
of nucleotides at a particular location.
• Silent Mutations.
• Not-So-Silent Mutations.
– Missense
– Nonsense
Substitution Mutations
• Original Sequence:
• Silent Mutation:
• Missense Mutation:
• Nonsense Mutation:
Point Mutations
• Insertions & Deletions – adding or
removing bases along the sequence.
– Results in a FRAME SHIFT because all other
codons “downstream” are affected.
– Can reverse if there are 3 insertions or 3
deletions.
Insertions & Deletions
• Original Sequence:
• Insertion/nonsense:
• Deletion/missense:
• 3-Nucleotide
Insertion/Deletion
(missense):
Assignment:
• Review chapters 16 & 17!
Transcription:
http://www.ncc.gmu.edu/dna/mRNAanim.htm
http://www-class.unl.edu/biochem/gp2/m_biology/animation/gene/gene_a2.html
Translation:
http://www.ncc.gmu.edu/dna/ANIMPROT.htm
http://www-class.unl.edu/biochem/gp2/m_biology/animation/gene/gene_a3.html
Overall:
http://www.johnkyrk.com/DNAtranslation.html