Genetic Code

• Codons composed of three nucleotides in RNA• Codon specifies amino acid or stop• Genetic code is redundant

Genetic Code Table

Reading Frames

• One of three possible reading frames contains protein message

Mutations In Protein-Coding Sequences

Missense Change to codon for different amino acid

Silent Change to codon for same amino acid

Nonsense Change to stop codon

Insertion/ Usually disrupt reading framedeletion

• Adaptors for associating codons with amino acids• Cloverleaf structure• Anticodon recognizes codon by complementary

base-pairing

Transfer RNA

Wobble Base-pairing

• Relaxed base-pairing at third position of codon

• tRNA may recognize multiple codons

Modified Nucleotides In tRNA

• post-transcriptional modifications

• Different synthetase for each amino acid• Amino acid attached to tRNA by high

energy ester bond

Aminoacyl-tRNA Synthetases

Sequential Action Of Adaptors

Editing By Synthetases

• Active site may mistakenly bind related amino acid• Incorrect amino acid is hydrolyzed at editing site

Growth Of Polypeptide

•Stepwise growth from N-terminus to C-terminus

• Large and small subunits• Composed of rRNA and proteins

Ribosome Structure

Translation On Ribosome

• P site contains tRNA attached to growing polypeptide

• A site binds incoming aminoacyl-tRNA

• Peptide bond formation by peptidyl transferase of ribosome

• Translocation of ribosome

• EF-Tu:GTP interacts with A site

• Codon-anticodon interaction leads to hydrolysis of GTP and dissociation of EF-Tu

• EF-G promotes ribosome translocation

Elongation Factors

Diphtheria Toxin

• Transfer of adenosine diphosphate ribose to EF-2

• Inhibits EF-2, which impairs ribosome translocation

Initiation In Eucaryotes

• Methionine-linked initiator tRNA to small ribosome P site (eIF-2)

• Load onto mRNA 5’ end (eIF-4E, eIF-4G, polyA)

• Scan for AUG• Assemble large

ribosomal subunit

Initiation In Bacteria

• Formylmethionine-linked initiator tRNA• Small ribosome subunit binds to Shine-Dalgarno sequence• Polycistronic mRNAs

Termination

• Stop codons not recognized by tRNA• Release factors bind to A site• Addition of H2O to peptidyl-tRNA

Polyribosomes

• Multiple ribosomes translating same mRNA

• Interaction of mRNA 5’ and 3’ ends

Selenocysteine

• Selenocysteine tRNA is charged with serine that is subsequently converted

• Encoded by UGA codon followed by special signal

Antibiotics

Tetracycline blocks binding of aminoacyl- tRNA to A-site of ribosome

Streptomycin prevents the transition from initiation complex to chain-elongating

ribosome; causes miscodingChloramphenicol blocks the peptidyl transferase reaction on ribosomesErythromycin blocks the translocation reaction on

ribosomesRifamycin blocks initiation of RNA chains by

binding to RNA polymerase

Folding During Synthesis

• Individual domains folded rapidly after their synthesis

Creating Functional Proteins

• Association with cofactors, proteins

• Covalent modifications

Molecular Chaperones

• Prevent inappropriate aggregation during folding• Recognize exposed hydrophobic regions• Hydrolyze ATP• Heat shock proteins

Hsp70 Chaperones

• Act early• Repeated cycles of target binding & release

mediated by ATP binding & hydrolysis

Hsp60 Chaperones

• Chaperonins• Isolation chambers• Cycles of target confinement & release

mediated by ATP binding & hydrolysis

Protein Quality Control

• Exposed hydrophobic regions indicate misfolding• Selective degradation of proteins that cannot be

correctly folded

Proteasome

• Protease activity in interior of cylinder

• Caps function in ATP-dependent unfolding and as gates for selective entry

Attachment Of Ubiquitin

• Attached to lysine on target; multiubiquitin chains• Ubiquitin attaches to E1• Transferred to E2 of ubiquitin ligase• E3 of ubiquitin ligase recognizes degradation signal

Regulated Degradation

• Modification of an E3 protein

• Modification of a target protein

Disease From Protein Aggregation

• Aggregates of misfolded proteins

• Cross-beta filaments• Neurodegeneration-

Huntington’s, Alzheimer’s, Prion diseases (Creutzfeldt-Jacob, bovine spongiform encephalopathy)