Chapter 24

Protein Synthesis and Controlled Protein Breakdown

Chapter Objectives

• Know the process of protein synthesis and the genetic code (don’t memorize the genetic code)

• Know how ribosomes work– Initiation, elongation, termination

• Know the key steps in protein folding• Understand how protein translation is

controlled• Know the process of protein degradation

The Genetic Code• Nonsense

mutation – stop codon

• Degenerate code – more than one codon per amino acid

• Conservation

Overview of Protein Synthesis

1. Amino acids are activated by aminoacyl synthetases using ATP– Amino acids are added to tRNA

2. tRNA molecules with their attached amino acid are brought to ribosome

3. The growing peptide chain is added to each new amino acid

4. Energy cost = 1 ATP & 2 GTP

tRNA

There are not 61 different tRNA molecules

tRNA Wobble

• Wobble allows for less tRNA molecules to be needed

• Notice orientation!!

Amino Acid Attachment to tRNA

• Must attach correct amino acid to tRNA– Wrong amino acid will be incorporated into protein

otherwise• Specific aminoacyl-tRNA synthetase for each tRNA /

amino acid pair– Need 20!

• No consistent recognition – Sometimes anticodon– Sometimes other regions of tRNA– Always side chain of amino acid

• Process driven by ATP hydrolysis

Specificity of Amino Acid Priming

• Prokaryote ribosomes– Subunits• 23S and 5S pieces of RNA

and 34 proteins• 16 S piece of RNA and 21

proteins

– Total• 50S (large subunit)• 30S (small subunit)

Ribosomes

http://www.mrc-lmb.cam.ac.uk/ribo/homepage/mov_and_overview.html

Translation Initiation

Shine-Dalgarno

30S ribosome subunit binds to Shine-Dalgarno sequence placing AUG in P site

Special met-tRNA recognizes IF2 (initiation protein factor 2)

IF3 prevents binding of 50S subunit prematurely

50S subunit then associatesIF2&3 are displacedGTP is hydrolyzed

Elongation

TerminationThis picture is from eukaryotes

Think of 60S as 50S and 40S as 30S

RF1 and 3 are RF1 and 2 in prokaryotes

Still need to bring in IF3 to prevent interaction of 30S with 50S

Overview

http://www.mrc-lmb.cam.ac.uk/ribo/homepage/mov_and_overview.html

Better overview

http://www.mrc-lmb.cam.ac.uk/ribo/homepage/mov_and_overview.html

Drugs that inhibit Translation

• Chloramphenicol – peptidyl transferase• Erythromycin – 50S inhibits translocation• Kirromycin or fusidic acid – prevents EF-Tu

release• Sparsomycin –peptidyl transferase inhibitor• Streptomycin – initiation misread• Tetracyclin – inhibits tRNA from binding

ribosome

Riboswitches

Selenocysteine

• Not a standard amino acid• Made from cysteine and selenium (toxic)• Uses UGA stop codon with special tRNA and

EF• Rare

Protein Folding

• Chaperones– ATP dependant– Heat Shock proteins

• Protein Disulfide Isomerases (PDI)• Peptidyl proline isomerase (PPI)

GroEL GroES chaperone

Prion Proteins

Programmed Protein Death

• Proteasome• Ubiquitin– Poly– Mono

Ubiquitin Pathway

• E1 is ubiquitin activating enzyme (ATP needed• E2 accepts Ub• E3 ligase activity

• Multiple E2/E3 combos