Protein Synthesis Flashcards
start codon
AUG
stop codons
UGA, UAA, UAG
direction of protein synthesis
NH2 -> COOH; read 5’ to 3’
silent mutations
point mutation that does not alter amino acid
missense mutation
point mutation that results in amino acid substitution
nonsense mutation
point mutation that results in premature stop codon
frameshift mutation
insertion or deletion (1 or 2 bases) that alters the reading frame
tRNA
contains the anticodon to base pair with mRNA codon
DHU loop
site on tRNA that binds tRNA synthetase
tRNA synthetase
responsible for forming bond between codon and anticodon
anticodon loop
part of tRNA that pairs with mRNA codon
3’ acceptor site
region on tRNA that is where amino acid attaches
anticodon of tRNA base
will be the amino acid identity in translation, no matter what amino acid is attached to the tRNA
Wobble pairing
weak interaction between 3rd codon and first anticodon
two important consequences of aminoacylation
energetics and fidelity
energetics in aminoacylation
hydrolysis of the ester linkage between amino acid attachment site and tRNA to allow new amino acid attachment is energetically favorable (releases phosphates)
aminoacylation fidelity
affinity for specific amino acids and proofreading by use of acylation and hydrolytic sites (cleaves incorrect amino acids)
Valyl-tRNA synthetase
binding pocket that exclude isoleucine based on size and hydrolytic sites
each amino acid has _
one tRNA synthetase that couples to it
tRNA synthetase reaction
hydrolysis of two ATP phosphate bonds –> energy for protein synthesis stored in tRNA-amino acid bond
bacteria ribosome
30S + 50S
eukaryotic ribosome
40S + 60S
small ribosome subunit
interacts with mRNA and translation factors
large ribosome subunit
23S/28S is a ribozyme (peptidyl transferase)
translation start site (bacteria)
determined by the Shine-Dalgarno sequence
translation start site (eukaryotes)
eIF4F scans mRNA to find AUG, additional factors needed
start site selection
mRNA, IF1, and IF3 and small ribosomal subunit associate
IF2/eIF2
in the GTP-bound form, bind to charged initiator tRNA and to pre-initiation complex
GTP hydrolysis by IF2/eIF2
stimulated by conformation of pre-initiation complex when codon:anticodon interaction is correct –> one GTP phosphate bond is cleaved for initiation
completion of initiation
large ribosomal unit binds
Shine-Dalgarno sequence
purine-rich region about 10 basepairs ahead of AUG
steps in initiation of translation (prokaryotes)
IF1 binds to 30S and blocks A site –> IF3 binds prevents 50S from binding –> Shine-Dalgarno binds and positions AUG at P site –> IF2-GTP brings fMet-tRNA to P site to pair with AUG codon –> IF2 hydrolyzes GTP to GDP releasing IFs and promoting 50S binding
GEFs
exchange GTP for GDP on GTPases
IF2 will ONLY bind_
fMet-tRNA
eIF2 (eukaryotes)
facilitates binding of initiating tRNA to 40S
eIF4A
RNA helicase to unwind
eIF4B
scans unwound mRNA to locate first AUG
eIF4E
binds to 5’ cap of mRNA
eIF2B
guanine exchange factor for eIF2
protein kinase R (PKR)
activated by dsRNA or interferons; can phosphorylate eIF2, causing it to bind GEF more tightly –> results in less active eIF2, slowing protein synthesis to decrease replication of virus
translation elongation (bacteria)
EF-Tu GTPase brings in aminoacyl-tRNA to A site –> GTP hydrolysis –> EF-Tu-GDP will now release tRNA –> peptidyl transferase binds fMet in P site with new amino acid in A site –> EF-G causes translocation
EF-G
translocase
EF-Ts
GEF for EF-Tu
eukaryote elongation factors
eEF1-alpha is EF-Tu and eEF1-beta is EF-Ts; eEF2 is EF-G
diphtheria toxin
targets eEF2 –> catalyzes ADP-ribosylation of eEF2 that blocks protein synthesis
translation termination (bacteria)
RF1 and RF2 recognize the stop codons –> RF3 mediates release of ribosomal subunits
eukaryote release factors
eRF1 binds termination codons and eRF3 promotes release
puromycin
