Ch 6B Notes

• The right strand
• the correct reading frame
• 3 potential reading frames for every segment of DNA

• the way the nucleotides are read from 5' to 3'
• the same RNA sequence can specify three completely different amino acid sequences, depending on the reading frame. But, only one contains the actual message

tRNA is constructed so that they require accurate base pairing only at the first two positions of the codon and can tolerate a mismatch at the third position

ester bond

activated

The initial check is via the aminoacyl-tRA synthetase, which couples a particular amino acid to its corresponding tRNA

THe second is the tRNA molecule itself, whose anticodon forms base pairs with the apprpriate codon on the mRNA

Through hydrolytic editing of incorrectly attached amino acids, the correct amino acid is rejected by the editing site

• 18S in euk
• 16S in pro

• large: forms peptidyl bond
• small: grabs onto mRNA

causes hybrid sites to form

it is a triple check system to ensure the correct amino acid is being added

1)Introduces a slight delay to the process

Coupling the GTP hydrolysis-driven changes in the elongation factors to transitions between different states of the ribosome speeds up protein synthesis

Specifically, EF-Tu binds GTP and aminoacyl-tRNAs simultaneously.

The dissociation of the inorganic phosphate group, which follows the reaction GTPàGDP and Pi causes a shift of a few tenths of a nanometer at the GTP-binding site

b. This tiny movement causes a conformational change to propagate along a crucial piece of alpha helix, called the switch helix, in the Ras-like domain of the protein.

c. The switch helix seems to serve as a latch that adheres to a specific site in another domain of the molecule, holding the protien in a “shut” conformation.d

. The conformational change triggered by GTP hydrolysis causes the switch helix to detach, allowing separate domains of the protein to swing apart, through a distance of about 4 nm.e. This releases the bound tRNA molecule, allowing its attached amino acid to be used.

• We ned key elements:
• - mRNA with 5' cap and poly-A tail
• - initiation complex: small ribosomal subunit/ initiator tRNA

initiator tRNA moves along RNA, searching for the first AUG

the initiator tRNA is bound to an eIF2 with GTP bound. The GTP and eIF2 dissociate once AUG is found

a specialized tRNA is charged with serine by the normal seryl-tRNA synthetase, and the serine is subsequently converted enzymatically to selenocysteine.

A specific RNA structure in the mRNA (a stem and lop structure with a specific nucleotide sequence) signals that selenocysteine is to be inserted at the neighboring UGA codon

This requires the participation of a selenocysteine-specific translation factor

it binds to newly formed backbone at an exon exon junction

mRNA, as it exits the nucleus, is test translated, removing EJCs from the mRNA

it causes a pre-mRNA to introduce a premature stop codon into the reading frame for the protein

nonsense-mediated mRNA decay: it destroys the abnormal mRNAs

if an in-frame stop codon is encountered before the final exon junction complex is reached, the mRNA undergoes nonsense-mediated decay, which is triggered by Upf proteins that bind to each EJC

it is more open and less highly ordered than the final folded form of the protein

the molten globule contains most of the secondary structure of the final form

they detect improper folding because of their exposed hydrophobic regions

act early, recognizing hydrophobic amino acids on a protein's surface

Aided by Hsp40 proteins, ATP-bound Hsp70 molecules grasp their target protein and hydrolyze ATP to ADp, undergoing conformational changes that cause the Hsp70 molecules to associate even more tightly with the target. 

Rapid rebinding of ATP indues dissociation of the Hsp70

misfolded protein captured

subsequent binding of ATP plus a protein cap increases the diameter of the barrel rim, which may stretch the client protein and confines it to an enclosed space

ATP hydolysis then occurs--> weakens complex

ATP binding then ejects the protein and the cycle repeats

tagged with a 76 amino acid tag called ubiquitin, which is attached to specific regions of proteins or subunits called degroms, which contain a lysine

proteins that make up the ring structure in the proteasome cap belong to this class of unfoldases called AAA proteins'

six subunits with ATP binding sites

causes shuttling in of substrate

3 component enzyme system

E1: Ub activating protein (generic finds Ub in cell, grabs ATP and uses ATP)

E2 and E3 are more specific; have to recognize different degrons

E2: Ub conjugating protein (transfers to a cysteine residue in E2)

E3: Ub ligase, but doesnt work without E2

has to be complementary

once bound, E2 transfers Ub to lysine residue and target protein

degron binds to binding site in E3

1) phosphorylation by protein kinase (In E2-P= off/ in E3-P= off)

2) allosteric transition caused by ligand binding 

3) allosteric transition caused by protein subunit addition

• protein subunit blocks degron
• Phosphorylation by protein kinase

unmasking by protein dissociation

creation of destabilizing N-terminus

the transcription level, but there are also other checks