Download E-site

ICE 4 Review
Translation and Gene regulation
Marty and Aaron
October 25th, 2014
Translation
From genes to proteins
Two enzymes involved in translation
amino acyl-tRNA synthetases
1. Adds amino acids to the 3’ of a tRNA
2. Proofreads that appropriate tRNA is
attached
ribosome
1. Site where tRNAs act as adaptors
between mRNA codons and amino
acids
2. Proofreads codon-anticodon pairing
3. Catalyzes peptidyl-transferase reaction
aa-tRNA synthetases
Adds amino acids
to 3’ adenine of tRNA
Cleaves amino acids
off of 3’ adenine of tRNA
Both steps catalyzed at the synthesis site:
Ribosomal Elongation
• A- site: where amino-acylated tRNAs are
deposited by EF-Tu when appropriate
codon-anticodon pairing occurs
• P-site: where the tRNA is bound to a
polypeptide as opposed to a single amino
acid
• E-site: where a spent tRNA goes before
getting kicked out
Peptide bond formation
Lone pair of electrons from the
amino group nitrogen in A attacks
carboxyl carbon in P.
Elongation Factors (“EF”s)
• We learned about two elongation factors used by the ribosome, both are
enzymes that hydrolyze GTP (i.e., “GTPases”)
• EF-Tu uses GTP/GDP to switch between two conformations:
• When bound to GTP, EF-Tu tightly binds a aa-tRNA
• When bound to GDP, EF-Tu binds aa-tRNA much more weakly
• This is why EF-Tu has to hydrolyze GTP to release an aa-tRNA into the ribosome’s
A-site
• EF-G uses the energy released by hydrolyzing GTP to push the tRNAs and
the mRNA in the ribosome over by one codon
Elongation Factor (EF)-Tu
deposits aa-tRNAs into the A-site
Elongation Factor (EF)-G translocates the
ribosome
after the
peptidyltransferase
reaction
Transcription/translation question
a) How many amino acids does this DNA sequence code for?
7
b) How many amino acids does this sequence code for if C18 is mutated to G?
4
c) How many amino acids does this sequence code for if G20 is deleted?
5
Transcription/translation question
3’
a) Please write the first 18 bases of the mRNA that would be transcribed, 5’ to 3’.
5’-AGGUCCACGGGUCCACCG-3’
b) What are the first five amino acids of the protein derived from the mRNA (please denote the N- and C-terminal
ends of the peptide) N-Met- Thr- His- Ile- Val- C
(ATG) (ACT) (CAC) (ATC) (GTT)
c) What are the last five amino acids of the protein derived from the mRNA (please denote the N- and C-terminal
ends of the peptide)
N-Ser- Gly- Ile- Gln- His- C
(AGC) (GGC) (ATC) (CAG) (CAT) (TAA = stop)
Mechanistic Ribosome Question from a pset
A
P
E
EF-Tu will hydrolyze a GTP and deposit an aa-tRNA in the A site.
codon: 5’-GUG-3’  valine
codon: 5’-GAU-3’  aspartic acid
Given the progress of the ribosome along the
mRNA diagrammed below, label the 5’ and 3’ ends
of the mRNA along with the A-, P-, and E-sites
5’
E P AE
3’
Label whether the N- or Cterminus of the growing
polypeptide chain is emerging
from the ribosome
N-terminus