1. Protein Biosynthesis Away from the Ribosome Genetic code Charging tRNA Ribosomes On the Ribosome Initiation complex Elongation factors Peptide bond formation Termination

2. Characteristics of the Code non-overlapping degenerate triplet basically universal over all living species polar no punctuation subject to miscues defines a reading frame

3. OLD MAN AND THE SEA Ernest Hemmingway Insert A OLD MAA NAN DTH ESE A OLD MAA ANA NDT HES EA OLD MAA ANA AND THE SEA OLD MAA NAD THE SEA Insert A A Insert AAA Insert A delete N N ^ Framing the Code Garbled In frame Garbled

4. tRNA

5. Charging tRNA Q: What is meant by Charging A: Charging means placing an amino acid on the 3’ (acceptor) end of the tRNA Q: So, what’s the big deal? A: There are 20 amino acids; the code is degenerate There could be 4 “isoaccepting tRNAs” competing for one Q: I still don’t see a problem A: One enzyme must recognize 4 different tRNA species and select the correct amino acid.

6. Q: One enzyme does all that? A: No, each tRNA has its own enzyme Q: What is this enzyme called? A: Its call Aminoacyl-tRNA Synthetase Q: So, there are 20 of these enzymes A: Yes Q: That makes the job a recognition a little easier then? A: Yes, but the enzymes still have to distinguish between look-alikes such as leucine and valine, glutamine and glutamate, tyrosine and phenylalanine.

7. Q: Are all aminoacyl-tRNA synthetases alike? A: Yes and no. Yes, they perform the same function, i.e., to recognize and transfer the correct amino acid to tRNA. Q: Why no? A: Because one class (Class I) looks for the anticodon on the tRNA, the other (Class II) looks for other features. Q: What else? A: Class I puts the amino acid on the 2’ position of the terminal ribose on tRNA, Class II only the 3’.

8. Q: So, how does aminoacyl-tRNA synthetase discriminate amino acids and different tRNA species? A: The key lies in the tRNA itself. Besides the anticodon, tRNAs have other bases that set them apart. These bases called “identity elements” are found in the terminal ends (acceptor stem) and internal in the tRNA. Q: Do they also proofreading? A: Yes, but sparingly Q: How sparingly? A: Enough to keep errors down to isoleucine mistaken for a valine once every 50,000 times. Ile-tRNA synthetase actually hydrolyzes the valine-AMP precursor.

9. Reaction: CH 3 CH 2 -CH 2 CH-COO - NH 3 + + ATP CH 3 CH 2 -CH 2 CH-C NH 3 + ~ O O-P-O-CH 2 O Ad OHHO O O CH 3 CH 2 CH-COO - NH 3 + L-Valine Enzyme Bound tRNA L-Leucine CH 3 CH 3 CH 2 CH-COO - NH 3 + CH 2 L-Isoleucine PP i

10. Codon-Anticodon Interactions Polarity 5’3’ 5’ Codon on mRNA Anticodon on tRNA Anticodon loop 3’ 5’ C G I (C, U) GCA mRNA 5’3’ Wobble base on anticodon 3rd position mRNAs are always read 5’ to 3’. mRNAs are always read 5’ to 3’. Alanine

11. Ribonucleoprotein Particles Ribosomes: The Staging Areas of Protein Synthesis 30S (40S) 16S RNA (18S) 23 Peptides (33) 50S (60S) 70S (80S) Monosomes 23S RNA (28S) 31 Peptides (49) 5S RNA (5S + 5.8S) * Mammalian 50S 30S tRNA sites mRNA Crevice

12. Polysomes Groups of ribosomes attached to a single mRNA