Exam 4: Protein Translation & post-translation Flashcards
5’ UTR
untranslated region at 5’ end of mRNA
3’ UTR
untranslated region at 3’ end of mRNA
open reading frame
area of mRNA translated into protein
monocistronic mRNA
mRNA contains only one open reading frame
polycistronic mRNA
mRNA contains several open reading frames
tRNA
anneal to 3-base codons on mRNA at 3-base anticodon region
have an amino-acid linked at the acceptor terminus
Protein synthesis occurs in
the cytosol, on ribosomes
Ribosomes
have two subunits, composed of 1-3 RNA molecules (rRNA) and dozens of proteins
Two types of ribosomes in humans
Cytoplasmic - synthesis of bulk of proteins
Mitochondrial - protein synthesis inside mitochondria
Translation initiation
IF2a activated by binding to GTP, binds to methionine-tRNA to form ternary complex
Terneray complex binds to a small ribosomal subunit
An mRNA molecule binds to structure to form pre-initiation complex
Pre-initiation complex binds to large ribosomal subunit to form initiation complex
eIF2a-GTP is hydrolyzed and GDP-eIF2a is released
eukaryotic initiation factor 2a (eIF2a)
activates initiation of protein translation by binding to GTP
Ternary complex
GTP-eIF2a and methionine-tRNA
Pre-initiation complex
ternary complex, small ribosomal subunit, and mRNA
Initiation complex
small & large ribosomal subunit, mRNA, tRNA-methionine
Elongation phase of translation starts after
the initiator methionine-tRNA binds to the P site of the ribosome
elongation factor (EF-1)
required to add a second tRNA to A site of ribosome
eukaryotic release factor (eRF)
pairs with stop codon
when attached GTP is hydrolyzed, peptide is released from P site and ribosome separates
Streptomycin
antibiotic - binds to small subunit and inhibits initiation, causes mistranslation of codons
Neomycin and gentamicin
antibiotic - bind to ribosomes and cause mistranslation of codons
Tetracycline
antibiotic - blocks A site of ribosomes and prevents tRNA binding
Chloramphenicol
antibiotic - prevents peptidyl bond formation in protein translation
Ricin toxin
removes adenine bases from various positions of the rRNA in large subunit, prevents translation
Diphtheria toxin
inactivates EF-2 by ADP-ribosylation, prevents translation
EF-2 (elongation factor)
provides energy for movement of mRNA one codon further, opening A site of ribosome
Methods for regulating translation
- Preventing the recognition of a start codon - binding protein to 5’ UTR of mRNA prevents recognition of start codon
- Regulating activity of initiation factors - phosphorylation of eIF-2 in response to certain stimuli inactivates initiation factor and turns off translation
Heat shock proteins (HSPs)
chaperone proteins that repair proteins damaged by heat and other stresses - help proteins fold correctly
Proteins destined for export from the cell are synthesized in the
endoplasmic reticulum
signal sequence peptide emerges from ribosome & recognized by SRP, moving the ribosome complex to the ER - binds to docking protein that transfers ribosome to transmembrane channel - translocon
As translation continues, peptide is threaded into ER
signal recognition particle (SRP)
recognizes signal sequence that peptide is being transported out of cell and binds to docking protein - transferring ribosome to translocon
translocon
transmembrane channel that threads newly synthesized proteins from ribosome into ER
Unfolded protein response
triggered by accumulation of unfolded proteins in ER - such as during starvation and cholesterol overload
Inhibition of global protein translation
Induce chaperone production - improve chances of proper folding
Considers apoptosis - if unfolded protein amount exceeds capacity for repair
Glycosylation is important because
- it changes the physical properties of the protein
- carbohydrates on protein surface are recognition sites for trafficking, protein interactions, recognition, and immune response
Glycosyltransferases
transfer sugar from an activated sugar nucleotide to an acceptor substrate
Specific for sugar donor, acceptor molecule, and type of bond formed
N-linked Glycosylation
Starts in ER before folding done, continues as protein goes through Golgi apparatus
oligosaccharide molecule is added to amino group of an asparagine residue
Modifications in Golgi yield what two types of N-glycosylated proteins?
High mannose type Complex type (mannose and 5 other charbohydrates)
O-linked Glycosylation
occurs only on fully folded proteins after it has reaced the Golgi aparatus
transfer N-acetyl-galactosamine to hydroxyl group of serine or threonine resideues on surface of protein
O-linked glycosylation is important in
blood types - attaches blood group antigens on red blood cells
H-antigen discriminates between self and foreign particles in blood - has 2 alleles, A & B
A allele
adds N-acetylgalactosamine to H antigen
B allele
adds galactose to H antigen
O allele
non-functional and adds nothing to H antigen
A & B alleles
express both forms of H antigen on surface of blood cells
Failure to hydroxylate proline
collagen disorders - Scurvy, Ehlers-Danlos syndrome, etc
failure to convert cysteine to formylglycine causes
multiple sulfatase deficiency (MSD) - sulfated glycosaminoglycans accumulate in lysosome
Timming at N-terminus of protein
proteolysis removes N-terminal amino acids & modifies new terminus - many proteins need a different start than methionine
Addition of hydrophobic moieties to protein
link membrane proteins to long-chain, hydrophobic molecules to change their surface properties
C-terminus additions
glycosylphosphatidyl-inositol (GPI) anchor to C-terminus to tether to external side of plasma membrane
Targeting of proteins to lysosome by
phosphorylating mannose residues of high-mannose glycoproteins
Mannose-6-P on protein surface targets vesicles to lysosome
Key features of proteins targeted for mitochondria
Unfolded - stabilized by chaperones
Synthesized with large N-terminal presequence - interacts with receptor in outer mitochondrial membrane (cleaved off by matrix proteases in mitochondria)
TOMs & TIMs
translocases of outer and inner mitochondrial membrane - channel through which preprotein enters mitochondrial matrix
Deafness-dystonia syndrome
mitochondrial disorder caused by mutation in a TIM component - impairs cellular energy production by preventing assembly of fully functional mitochondria
Cystic fibrosis
caused by deletion of one codon from CFTR1 gene
Interferes with folding and glycosylation of protein
CFTR protein is moved into cytosol and degraded instead of being sent to plasma membrane
I-cell disease
transfer of phosphate to mannose is impaired
lysosomal proteins do not reach their compartment & function is compromised - leads to accumulation of undegraded proteins in lysosomes
In fibroblast - dense bodies of nonfunctional lysosomes & content
In serum - lysosomal proteins that did not reach lysosomes
Mechanisms for protein degradation in cell
Lysosome & proteasome
Lysosome
nonspecific degradation of extracellular and intracellular proteins
can digest itself - autophagy
contains high concentrations of diverse hydrolytic enzymes
Proteasomes
required for specific degradation of cytoplasmic proteins
Ubiquitin
mark protein for proteasomal destruction - needs multiple units of ubiquitin on same protein (poly-ubiquination) to signal for destruction
Activated by E1 enzymes, conjucated to E2 enzymes, and ligated to targets by E3 proteins
Recycled - not degraded by proteasome with protein
Enzyme E3
identifies proteins for ubiquination & transfers ubiquitin to protein
Specific isozymes for certain classes of substrates
Protein lifespan determined by:
Conformation (correct folding, no hydrophobic domains on surface of protein)
N-terminal residue (arginines or lysines are less stable than proteins with methionine or serine at N-terminal)
Other sequence elements (PEST sequences shorten lifespan of protein)
PEST sequences
proline-glutamine-serine-threonine sequence in protein
Shorten life of protein
