Download Announcements Chapter 15 Proteins: Function Proteins: Function

Announcements
• Lab Next Week
Chapter 15
• Help Session: Monday 6pm LSS 277
The Genetic Code and
Translation
• Office Hours
Proteins: Function
Proteins: Function
Luciferase
• Enzymes
• Transport
• Structural
Components
• Regulation
• Communication
• Defense
• Enzymes
• Transport
• Structural
Components
• Regulation
• Communication
• Defense
Ricin
Proteins: Structure
Fibroin
Peptide Bonds
Figure 15.6
• Composed of amino
acids
– 20 amino acids, similar
in basic structure
• Joined by peptide
bonds, forming
polypeptide chains.
1
Figure 15.7
The Genetic Code
• How many nucleotides are necessary for
amino acid specification?
• 20 amino acids
Protein Structure
The Genetic Code
• How many nucleotides are necessary for
amino acid specification?
• 20 amino acids
• One?
– 4 bases (AGCU) = 4 possible codons
The Genetic Code
• How many nucleotides are necessary for
amino acid specification?
• 20 amino acids
• One?
• Two?
2
– 4 bases at two positions = 4 = 16 codons
The Genetic Code
• How many nucleotides are necessary for
amino acid specification?
• 20 amino acids
• One?
• Two?
• Three?
3
– 4 bases at three positions = 4 = 64 codons
The Genetic Code
• How many nucleotides are necessary for
amino acid specification?
• A triplet code is the most efficient way
to code for all 20 amino acids
• Shown by Crick et al in 1961
2
Cracking the Genetic Code
?
Cracking the Genetic Code
• Homopolymers: Poly (A), Poly (U), Poly
(G), Poly (C)
– Determine amino acids for UUU, AAA, GGG,
CCC
Nirenberg and Matthaei (1961)
Cracking the Genetic Code
The Poly (G)
results were
uninterpretable!
uninterpretable!
Phe
Pro
Figure 15.9
Lys
?
Cracking the Genetic Code
• Homopolymers: Poly (A), Poly (U), Poly
(G), Poly (C)
– Determine amino acids for UUU, AAA, GGG,
CCC
The Genetic Code
?
• By using other clever methods, the genetic
code was fully understood by 1968.
– First started investigating in 1961.
Figure 15.12
3
The Genetic Code
The Genetic Code:
Code Redundancy
Stop Codons
Also called
termination codons
or nonsense
codons
• Isoaccepting tRNAs carry the same
amino acid but have different anticodons.
• Codons that specify the same amino acid
are synonymous.
• Sense codons specify an amino acid
– 61 sense codons
• Only 20 amino acids.
– The genetic code is a degenerate code
Figure 15.12
Degenerate Code:
Code Amino acids may be specified by
more than one codon.
Degenerate?
Figure 15.13
Wobble:
Wobble “Flexibility” in the pairing of the 5’ base of the
anticodon with the 3’ base of the codon
Initiation Codons and Reading
Frame
• Initiation codon:
codon AUG
– Bacteria: specifies N-formylmethionine
– Eukaryotes: specifies methionine
• Genetic Code is nonnon-overlapping
– Except in some viruses
• 3 possible reading frames
4
Figure 15.14
The Genetic Code is
Universal**!
3 different
reading
frames
**The (Almost
Almost) Universal Genetic Code
The problem set
frustrates and and
infuriates me!
4 Stages of Translation
The Process of Translation
• 1) tRNA charging:
charging
– Amino Acids bind to tRNA
• 2) Initiation:
– Necessary components bind to ribosome
• 3) Elongation:
– Amino acids joined to growing polypeptide
• 4) Termination:
– Protein synthesis stops at stop codon, translation
components released from ribosome
5
Stage 1:
1 Binding of AA to tRNA
• Aminoacyl-tRNA synthetases:
– 20 different synthetases
– Each recognizes a particular amino acid
• Based on size, charge, R group
– Each recognizes all the tRNAs associated
with its amino acid (isoaccepting tRNAs)
• Sequences in DHU arm, anticodon loop, acceptor
stem critical to tRNA recognition
Figure 15.15
Invariant Positions
Single Synthetase Recognition
Multiple Synthetase Recognition
tRNA Charging:
2 Step Process that
Requires energy in the
form of ATP
Stage 2:
2 Initiation
• All ingredients required for translation are
assembled:
Stage 2:
2 Initiation (Bacteria)
• All components required for translation are
assembled:
– mRNA
– Ribosome (small and large subunits)
– Initiation factors (3 proteins)
– Initiator tRNA with N-formylMethionine
attached (fMet-tRNAfMet)
– Guanosine triphosphate (GTP)
6
Initiation: Step 1
• mRNA binds to small subunit of
ribosome
– Initiation factor 3 (IF-3) keeps large and small
subunits separated during initiation
• Key consensus sequence in Bacteria for
ribosome binding: ShineShine-Delgarno
sequence
– Complementary to a sequence near 3’ end of
16S rRNA
Initiation: Step 1
• mRNA binds to small subunit of
ribosome
– Initiation factor 3 (IF-3) keeps large and small
subunits separated during initiation
• Key consensus sequence in Bacteria for
ribosome binding: ShineShine-Delgarno
sequence
– Complementary to a sequence near 3’ end of
Cool! An
16S rRNA
example of an
RNA-RNA
interaction!
Figure 15.16
Initiation: Step 1
IF-3 prevents
large subunit from
binding
Initiation: Step 2
Figure 15.16
• fMetfMet-tRNAfMet attaches to initiation
codon
– Facilitated by Initiation Factor 2 and GTP
– Initiation Factor 1 helps keep large and small
subunits apart
• 30S Initiation Complex
– Small ribosomal subunit, mRNA, fMettRNAfMet, GTP, Initiation Factors
Initiation: Step 2
Formation of the
30S Initiation
Complex
7
Initiation: Step 3
Figure 15.16
• 70 S Initiation Complex
– Large subunit of ribosome joins Initiation
Complex
– IF-1 and IF-2 depart
Initiation: Step 3
Formation of the
70S Initiation
Complex
Eukaryotic Initiation
Figure 15.21
• No Shine Delgarno sequence
– 5’ Cap important in ribosome-mRNA binding
• More initiation factors required
• Poly(A)
Poly(A) tail bound proteins
– Interact with 5’Cap bound proteins
8