L14: tRNA and codon anticodon recognition

messenger RNA

carries the genetic code to produce protein

ribosomal RNA

• structural component of ribosome

• major roles in ribosomal function and codon-anticodon recognition and transpeptidation

transfer RNA

• carries the aa to the ribosome

• converts codons into aa

• adapter molecule that matches codon to their aa

ends of tRNA

• one side recognizes the codon

• other side attaches to an aa

how do tRNAs align on mRNA

• align side by side to line up aa in correct order

• no more 2 are side by side at once

how do aa bind to tRNA

• aa covalently binds to 3’ end of tRNA

• called ‘charging the tRNA with aa’

aminoacyl-tRNA

• aa-tRNA

• tRNA when its attached to an aa

tRNA structure

• made of short, ssRNA

• 75 nucleotides long on average

• folds into an L shaped 3D structure via base pairings

• looks like a clover leaf in 2D

4 major structures of tRNA

• acceptor stem

• anticodon loop

• D-loop (close to 5’ end)

• TpsiC-loop (close to 3’ end)

acceptor stem

• made of the 5’ and 3’ ends of tRNA

• 3’ end sticks out with the terminal CCA sequence (conserved)

• aa gets covalently attached to 3’ end of the last adenosine

anticodon loop

• occurs on the opp side of acceptor stem

• contains the 3 anticodon bases that matches the codon

tRNA species on diff aa

• have diff surface shapes

• enzymes must tell them apart to charge tRNAs with correct aa

• tRNA bases are heavily modified to fine tune surface shape and function

where are codons and anticodons paired

inside ribosomes

what does ribosome do

• holds tRNA and mRNA at a very specific angle to make them interact in a controlled environment

• judges if codon and anticodon are correctly matches

where is the codon position 1

at mRNA 5’ end

where is the codon position 3

at mRNA 3’ end

where is the anticodon position 1

at tRNA 5’ end

where is the anticodon position 3

at tRNA 3’ end

codon-anticodon position pairings

• antiparallel

• codon 1 with anticodon 3

• codon 2 with anticodon 2

• codon 3 with anticodon 1

how to write anticodons in questions

5’ to 3’

wobble base pairing

• when ribosome matches codon to its anticodon, unconvential base pairing can occur

• AAG anticodon can match with either UUC (normal) or UUU

• both code of Phe

• structural basis for codon degeracy

wobble position

• codon position 1 and 2 follow normal base pairing

• codon position 3 and anticodon position 1 are the wobble positions

Inosine

• purine

• unconventional base

• anticodon position 1 can base pair with multiple bases

• can pair with C, A, U

wobble in eukaryotes vs bacteria

• eukaryotes are more stringent

• does not have as much wobble flexibility

what can bacterial U pair with

A,G,I

what can bacterial C pair with

G,I

what can bacterial A pair with

U,I

what can bacterial G pair with

C,U

what can eukaryotic U pair with

A,G,I

what can eukaryotic C pair with

G,I

what can eukaryotic A pair with

U

what can eukaryotic G pair with

C

where does an aa get charged

to the 3’ OH of the last ribose sugar

2 steps of tRNA charging

1. carboxyl end of aa gets attached to the alpha phosphate of ATP

2. aa is transferred from AMP to the 3’ end of the tRNA

what happens when aa attaches to the alpha phosphate during tRNA charging

• produces aa that is attached to an AMP

• 2 phosphodiester bonds of ATP broken to energize this reaction

AMP

adenylated amino acid

aminoacyl-tRNA synthethases

• aaRS

• enzymes that facilitate joining aa to tRNA

• a single aaRS responsible for putting one type of aa to all tRNAs that have the corresponding anticodon for that aa

• ensures that the correct aa gets charged to tRNA

how does aaRS ensure the correct aa gets attached to tRNA

• recognizes different shapes of the tRNA to find the correct one

• specifcally looks at the shapes of the anticodon loop and acceptor stems

• recognizes diff shapes and properties of aa

what does aaRS use to add the correct aa to tRNA

• 2 catalytic sites

• synthesis and editing site

synthesis site

• adds the aa to tRNA

• aa larger than the correct one cannot enter this site

• only the correct or a smaller aa gets added to tRNA

editing site

• removes any aa that is incorrectly joined to tRNA

• after an aa is added to tRNA, it leaves the synthesis site and tries to enter the editing site

• this site allows all aa to enter except the correct ones

• any aa that enters, gets removed

draw tRNA

x slide 9

draw tRNA in ribosome

x slide 11/12