Exam 4: Lecture 1 Flashcards
1
Q
Translation
A
-tasked with transferring mRNA transcript sequence into protein sequence
2
Q
Ribosome Subunits
A
- consists of small subunit and large subuinit
- each subunit consists of multiple different proteins
- both associated with non-coding RNAs (ncRNA)
3
Q
ncRNA
A
-aids in stabilizing interactions with mRNA transcript
4
Q
Translation Brief Process
A
- mRNA transcript bound by small ribosomal subunit
- initiator tRNA bound to modified Methionine (fMet) brought into contact with first AUG codon of coding sequence
- large ribosomal subunit added to complex
- complete ribosome reads transcript 5’ to 3’ and generates appropriate protein sequence
- process repeated till mRNA transcript is degraded
5
Q
Polysomes/Polyribosomes
A
- multiple ribosomes that bind each mRNA transcript
- increases amount of protein produced prior to degradation because multiple ribosomes are simultaneously translating mRNA
6
Q
Increasing Protein Production
A
- mRNA transcript organized into circular structure
- 5’ UTR and 5’ CAP structure bound initiation factors that interact with Poly-A Binding Protein which is bound to 3’ Poly-A tail
- circular conformation protects ends from degradation and can be translated by multiple ribosomes for longer period of time than linear mRNA (prefer to degrade single stranded DNA)
- eventually falls apart
7
Q
Ribosome Composition
A
- small and large subunit
- subunits composed of many individual proteins
8
Q
Centrifugation
A
- used in early attempts to understand ribosome structure
- intact ribosomes placed in sucrose solution and spun at high velocities
- size of protein complex directly proportional to sedimentation velocity
- i.e. intact ribosome travels faster through gradient than small or large individual subunits
- likewise large subunit sediments at higher velocity than smaller
- after isolated, further disrupted to yield individual proteins sequenced by Fred Sanger’s methods
9
Q
A, P, and E sites
A
- three channels for tRNA binding within ribosome
- initiator tRNA recruited to P site where it interacts with first AUG within coding region of mRNA transcript
- once initiator tRNA and AUG codon intact with P site, large subunit recruited
10
Q
Alignment
A
- Aligning first codon with P site automatically ensures second codon aligned with A site
- second tRNA enters ribosome and interacts with transcript via A site
- peptide bond forms between first two amino acids then ribosome moves to next codon.
- moves initiator tRNA into E pocket where it’s ejected
- second tRNA moves into P pocket leaving empty A pocket
- each subsequent tRNA enters ribosome through A site to interact with mRNA
11
Q
Translation Initiation Sequences
A
- ribosome must be recruited to 5’ end of mRNA transcript
- prokaryotes: Shine-Dalgarno sequence located 3-9 bases upstream first AUG
- eukaryotes: Kozak sequence encompasses few bases upstream and downstream AUG
12
Q
Binding to Initiation Sequences
A
- after binding to sequences AUG placed in P pocket
- non-coding ribosomal RNAs associated with small ribosomal subunit important in interacting with mRNA and for ensuring first AUG aligned with P pocket
13
Q
Initiation of Prokaryotic Translation: Blocks on Sites
A
-initiation factors bind to E and A sites to block initiator tRNA from entering
14
Q
Initiation of Prokaryotic Translation
A
- steps after blocking
- mRNA and initiator RNA recruited to small ribosomal subunit
- contact between ribosome and mRNA mediated by RNA:RNA interactions
- 16S ribosomal RNA base pairs with Shine-Dalgarno sequence in mRNA
- 5-9 base sequence that allows just enough space for AUG to be in P pocket which ensures AUG’s position
- large ribosomal subunit recruited to make complete ribosomal unit
- anticodon of initiator tRNA makes contact with AUG codon in P pocket begins elongation
- IF’s (1 and 3) removed from A and E pockets
15
Q
Length of Sequence for Alignment
A
- too long, AUG won’t line up in P pocket (may be in A pocket or completely out of ribosomal subunit)
- too short also causes problems
16
Q
Differences in Eukaryotic Translation
A
- initiator tRNA recruited to small ribosomal subunit independently of mRNA transcript
- recruitment of transcript does not ensure alignment of initiator
17
Q
Similarities between Prokaryotic and Eukaryotic Translation
A
- number of initation factors required to displace large ribosomal subunit and to block E and A channels
- leaves P channel only one open for tRNA
18
Q
Translation Elongation (study example in notes)
A
- subsequent tRNAs (after initiator) enter A pocket
- second codon enters and codes for amino acid
- tRNA attached to that protein recruited to A pocket where anticodon pairs with the codon
- Met transferred to tRNA containing the amino acid coded for and a peptide bond will form between the Met and other amino acid
- ribosome moves over slightly moving AUG codon into E pocket and the subsequent is in P pocket and third is in A pocket
- empty tRNA in E pocket expelled and A pocket is filled by tRNA that base pairs with codon
- process repeated again and again until ribosome reaches stop codon
- then dissociates from mRNA and nascent protein is released
19
Q
Eukaryotic Translation
A
- ready to interact with mRNA transcript when pre-initiation complex has formed
- 5’ end of mRNA bound by IFs that interact with IFs bound to small ribosomal subunit
- recruitment of mRNA to small ribosomal subunit does not result in alignment of initiator AUG in P pocket (difference)
- small ribosomal subunit moves along mRNA until it finds AUG which is recognized when anticodon of tRNA bases pairs with initiator codon
- initiator codon:anticodon binding stabilized, large ribosomal subunit recruited to make complete ribosome
- IF blocks expelled from E and A pockets and elongation begins
