Functions & Dysfuncions Of Protein Processing Flashcards
How does a codon recognize an amino acid?
Through genetic code
Genetic code
DNA (nucleotide sequence)—> protein (AA) using mRNA
Codon
Group of 3 nucleotides
What do codons code for?
61 triplet codons code for 20 Amino Acids or 3 stop translation codons
Characteristics of code
Degenerate (some AA are coded by multiple codons)
Standard, universal
Not punctuated
Non-overlapping
Silent mutation
Codon changes, AA is the same
Missense Mutation
New codon —> new amino acid
Can change protein or not
Nonsense mutation
Change in codon —> stop codon
Protein degrades or stops truncated
Frameshift mutation
One nucleotide is added/deleted it moves the entire sequence one over so AA sequence is disrupted; leads to nonfunctional protein
Sickle cell anemia
Due to missense mutation that changes GAG to GTG which changes Glutamic Acid (negatively, charged) to Valine (non polar, non charged)
Mutation causes the RBC to form a rigid, rod-like structure and deforms RBS-> clog capillaries b/c they cant carry O2
Duchenne Muscular Dystrophy
Frameshift mutations in dystrophin gene
(In-frame: mild form only truncated forms, out of frame: actual MD)—> little or no expression of the dystrophin gene —> muscle wasting
mRNA structure
5’ cap: 7-methylguanosine cap
3’ end: poly-A tail (lots of A amino acids)
coding region: codons for amino acids
tRNA structure
cloverleaf (secondary structure)
unpaired nucelotide regions: Anticodon loop (binds to codon on mRNA) & 3’ CCA terminal region
3’ region of tRNA
CCA region that binds the amino acid w the corresponding codon
Anticodon loop
3 nucleotides that pair w a complementary codon
Aminoacyl tRNA
tRNA that arrys the Amino Acid –> needs to be activated
Enzyme that activates Amino Acid
Aminoacyl tRNA synthetase
Aminoactyl tRNA synthetase
activate amino acids by serving as a second genetic code to maintain the fidelity of protein synthesis
Steps of Amino Acid Activation
using AMP to COOH end of amino acid (breaks the AMP) which gives energy to bind codon to the mRNA
Prokaryotic Ribosomes
Total: 70S
Small: 30s, Large: 50s
different structure so that antibiotics can target ONLY prokaryotic mechanism
eukaryotic ribosomes
Total: 80s
Small: 40s, Large: 60s
A site of ribosome
where the mRNA codon is exposed to receive the aminoacyl tRNA
***except Met tRNA (start codon)
P site of ribosome
where the aminoacyl tRNA is attached; holds the tRNA w the growing polypeptide chain
E site of ribosome
where the protein will exit the ribosome
Initiation of translation
small ribosomal sub unit binds to P site of SOMETHING where there is an EIF (EIF2) that is bound to GTP (source of energy). Large subunit attaches to form complex. Next tRNA comes in (based on sequence) and forms first peptide bond to methionine
Where are EIF4s attached?
Poly-A tail
What is the bond from one amino acid (in A site) to the growing chain (P site?) By which enzyme?
Peptide bond (CO-NH) by Peptidyl transferase
Elongation of Translation
purpose: links AA to growing polypeptide w elongation factors that play a a role in proofreading
aminoacyl tRNA is attached to GTP-bound elongation factor.
loading of AA: anticodon base pairs w codon on the A site
Termination of Translation
Triggered by stop codons: UAA, UAG, UGA which are recognized by RELASE factors (RFs)
RFs bind to the A site and cleaves the ester bond between the Carbon & the tRNA (reacts w water to make the COOH group and makes the protein)
peptide chain is released from ribosomal complex and then ribosome dissociates by GTP hydrolysis
Stop Codons
UAA, UAG, UGA
U Are Away, U are Gone, U Go Away
Polysomes
cluster of ribosomes that collectively make polypeptide efficiently
streptomycin
prokaryotic; binds to 30s to disrupt initiation
chloramphenicol
inhibits peptidyl transferase in the mitochondria of prokaryotes
clindamycin & erythromycin
prokaryotic; bind to 50s and blocks translocation of ribosome
erythromycin
treats purtussis
streptomycin
prokaryotic; binds to 30s and interfers with binding fmet-tRNA (first tRNA)–> interferes w joining 30s and 50s
cycloheximide
eukaryotic; inhibits peptidyl transferase
diphtheria
eukaryotic; inactives GTP-bound eEF-2 and interferes w ribosomal translocation
shiga toxin & ricin
eukaryotic; binds to 60s and blocks entry of aminoacyl-tRNA to ribosomal complex
puromycin
prokaryotic/eukaryotic causes premature chain termination; exact mechanism is unknown but somehow it enters the A site and adds to the growing chain w a puromyclated chain which results in early release
Cytoplasmic pathway
protein sorting pathway; for proteins that are destined to go to cytosol, mitochondria, nucleus and peroxisomes; translation begins & ends on free ribosomes; no signals –> stay in cytoplasm; those w signals will go to specific organelles w help of chaperone proteins (TrasnsporterInnerMembrane and TrasnporterOuterMembrane)
secretory pathway
protein sorting pathway for proteins destined for ER, lysosomes, plasma membrane or for secretion;
signal & pathway that sends to Mitochondria
cytoplasmic; N terminal hydrophobic alpha helix
signal & pathway that sends to nucleus
cytoplasmic; lysine and arginine rich
signal & pathway that sends to peroxisome
cytosplasmic; SKL sequence
signal & pathway for secretory proteins
secretory pathway; Trp-rich domain
signal & pathway that sends to lysosome
secretory pathway; Mannose 6 phopshate
signal & pathway that sends out of cell
secretory pathway; Stop tsrf
signal & pathway that sends to ER lumen
secretory pathway; lys-asp-glu-leu (KDEL)
how do proteins pass through the mitochondrial membrane?
TOM (outer) & TIM (inner) and protected by binding to heat shock protein 70 (HSP70)
Where does translation occur for secretory pathway?
translation begins on free ribosome but ends on ribosomes that are going to ER
Signal peptide for secretory pathway
all have ER targeting (15-60 AA at N terminus w 1 or 2 basic AAs near N term and hydrophobic on C side)
signal recognition particle (SRP)
binds to ER and ribosome, wraps itself around the ribosome mRNA peptide complex and stops translation for a brief sec (resumes when protein is directed into lumen)
how does the protein get released?
enzymes on luminal side cleave the signal and release it additional signals will tell it where to go
I-cell disease
due to: lack of mannose 6P enzyme
protein folding
small proteins spontaneously fold into naive conformations; large proteins need chaperones & chaperonins to ensure protection and proper folding
chaperonins
barrel shaped compartments that admit unfolded proteins and use ATP to help fold
proteolytic cleavage
converts inactive zymogen –> active enzyme
acetylation
adding a covalent bond to Amine (-NH3) on lysine; critical for gene regulation w histones (HAT/HDAC); patterns of histone modification are genetic
O-glycoslyation
adds hydroxy group to serine or threonine
n-glycosylation
acide-amide (-CONH2) to asparginine or glutamine; precursor sugar trasferred from phospho dolichol
phosphorylation
phosphate linked via creating ester, uses OH group to serine, tyrosine kinase, threonine and also asparatate and histidine; phosphate group is ultimately removed by phosphatase; seen in cell growth, proliferation, differentiation, oncogenesis
Disulfide bonds
inter/intra molecular disulfide (SH) bonds stablize proteins; bond between SH groups of two cysteines in ER lumen; enzyme: protein disulfide isomerase
modifications of collagen
modifications are important for assembly of collagen; utilizies lysyl hydroxylases + ascorbic acid (defects–> skin/bone/join disorders
ehlers-danlos snydrome
overly flexible joints due to lysyl hydroxlase defects
Alzheimer’s Disease: mechanism
when normal Amyloid precurose protein breaks down to form Amyloid beta peptide more than normal
AD: misfolding/aggregation of AB –>clumps/plaques
Also: hyperphosphorylation of Tau–> neurofibrillary tangles
both mutations in APP and Tau–> familal forms , aging–> sporadic forms
Parkinson’s
caused by: alhpha synuclein (AS) –> insoluble fibrils–> lewy bodies in dopamingergic neurons in substania nigra– > death of neurons–> reduced dopamine –> motor impariment
mutations of AS–> familial form, aging–> sporadic
Huntington’s
mutation in huntingtin gene–> expansion of GAG triplet polyglutamine repeats –> intramolecular hbonds are formed–> misfold and aggregate–> selective death in basal ganglia
Creutzfelt-Jacob Disease
cause: misfolding of prion proteins –> transmisslbe spongiform encephalopathies
danger: transmissible: can convert regular proteins to misfolded
