Functions & Dysfunctions of Protein Processing Flashcards
What are the start and stop codons?
AUG (Methionine); UAG,UAA, UGA
What are the ribosomal and eukaryotic subunits in translation?
Prokaryotes:
30S and 50S
Eukaryotes:
40S and 60S
What are the prokaryotic translation inhibitors?
1) Streptomycin - binds to 30s, impairs initiation
2) Clindamycin/Erythromycin: binds to 50s, blocks ribosome translocation
3) Tetracycline - binds to 30S, impairs elongation
4) Chloramphenicol - inhibits peptide transferase activity, impairs peptide bond formation
Eukaryotic Translation Inhibitors
1) Shiga toxin & Ricin - bind to 60s, blocks aminoacyl-tRNA
2) Diptheria toxin: no ribosomal translocation - inactivates GTP-bound EF-2
3) Cycloheximide - impairs peptide bond formation
What is an elongation Inhibitor?
Puromycin - premature chain termination
-Stops ribosome from functioning
What are the types of mutations (w/ clinical example)?
1) Silent - no AA change
2) Missense - changes AA in a protein with no impact on function (Sickle cell)
3) Nonsense - codon becomes stop codon - chain termination
4) Frameshift - one or more nucleotides deleted or inserted —- HUGE change in sequence (Duchenne Muscular Dystrophy)
Sickle Cell Anemia
- Missense mutation
- Glutamic acid changes to Valine
- HbA conformational change, RBC become rigid, rod-shaped
- RBCs = poor 02 capacity, clog capillaries
Duchenne Muscular Dystrophy
- Frameshift mutation on in-frame and out-of-frame Dystropin gene (Dystropin…think “dystrophy”)
- No expression of dystrophin
- In males, muscle wasting, death within 10 years
- Mild form = Becker muscular dystrophy
Protein Sorting Pathways
1) Cytoplasmic - cytosol, mitochondria, nucleus, peroxisomes. Starts and ends on free ribosomes
2) Secretory - ER, lysosomes, plasma membranes, secretion. Translation starts on free ribosomes, ends on ribosomes sent to the ER
Cytoplasmic Pathway destinations and translocation signals
1) Cytoplasm - None
2) Mitochondria - N-terminal hydrophobic alpha helix
3) Nucleus - KKKRK sequence
4) Peroxisomes - C-terminal SKL sequence
Secretory Pathway destinations and translocation signals
1) ER lumen - C-terminal KDEL
2) Lysosomes - Mannose 6 Phosphate (you know this)
3) Secretion - Tryptophan-rich domain
4) Membranes - N-terminal apolar region
Explain the Mitochondrial Protein Import process
1) Translation sequence recognized by TIM and TOM — inner and outer membrane proteins, respectively
2) Proteins moved across membrane
Explain the Nuclear Protein Import Process
- Proteins move through nuclear pores
- Large proteins need NUCLEAR LOCALIZATION SIGNALS
- Four basic residues (Lysine & Arginine)
What are the steps in the Secretory Pathway?
- Proteins have:
- ER signal peptide
- 15-60 AA at N-terminus
1) SRP (signal recognition particle) binds to ER signal and ribosome and then wraps around ribosome-mRNA complex
2) This halts translation and SRP guides/docks to translocon (protein receptor)
3) Once in ER lumen, translation resumes, enzymes cleave signal to release protein
4) Protein undergoes PTMs
What is I-Cell Disease?
- Deficiency in Mannose-6-Phosphate
- Proteins not target to lysosomes
- Results in:
1) Failure to Thrive
2) Developmental delays
3) Abnormal skeletal development, hepatomegaly
4) Death usual by age 7
Explain protein folding
- The idea that small proteins can spontaneously fold into tertiary conformations
- Large proteins can’t, need help from CHAPERONES
What are chaperones and chaperonins?
- Molecules that help large proteins fold into their tertiary structure without risk of:
1) aggregation
2) proteolysis
-Chaparonins are barrel-structure molecules that use ATP to fold large, unfolded proteins
Name a chaperone and chaperoning protein
Chaperone - HSP70 (heat shock protein)
Chaperonin - HSP60
Type of PTMs
1) Protein folding
2) Proteolytic cleavage
3) Covalent modifications
Explain Proteolytic cleavage
-Reveals active site of enzymes, making them active (i.e. trypsinogen -> trypsin)
What are some types of covalent modifications?
1) Glycosylation
2) Phosphorylation
3) Disulfide bond formation
4) Acetylation
Glycosylation (detail, functional group, residue affected) —– CLINICAL?????
1) 0-glycosylation; OH; Ser, Thr
2) N-glycosylation; Acid-Amide; Asn, Glu
-Cataract formation, similar to galacticol
Acetylation (detail, functional group, residue affected, impacts)
- Covalent amine linkage
- Amine
- Lysine
- Catalyzed by HAT and HDAC, important for GENE REGULATION
- Impacts are heritable (epigenetics)
Disulfide Bond formation (detail, functional group, residue affected, impacts)
- oxidation leading to cysteine residue linkage
- SH (sulfahydryl)
- Cys
-Stabilize proteins, formation occurs in ER lumen
Phosphorylation (detail, functional group, residue affected, impacts)
- Phosphate added via ester bond
- OH
- Ser, Try, Thr, Asp, His
-Important for enzyme activity, protein function, cell growth, cancer???
Explain the PTMs of collagen
- Collagen biggest structural protein
- Modification necessary for collagen
- Vitamin C = important for LYSYL and PROLYL HYDROXYLASES
- Defects = skin, bone, join problems —- Ehlers-Danlos
Ehlers-Danlos Syndrome
- From defect in Prolyl hydroxylases In collagen formation
- overly flexible joints
- blood vessel walls, intestines or uterus may rupture
Alzheimer’s Disease (AD)
- Memory loss, cognitive dysfunction as a result of:
1) amyloid precursor protein (APP) breakdown to amyloid beta peptide
2) Hyperphosphorylation of Tau
Brain aging = sporadic form
Parkinson’s disease (PD)
1) a-synuclein (AS) protein deposit as Lewy bodies in neurons (familial form)
2) Cause reduced availability of dopamine (from tyrosine precursor)
Brain aging = sporadic form
Huntington’s Disease (HD)
- Mutation in Huntington gene = CAG repeats
- Leads to abnormal Huntington protein, which misfiles and aggregates
- Cell death in basal ganglia
Creutzfeldt-Jakob Disease
- “Prion Protein” misfolding
- TRANSMISSIBLE, like catching a cold
- Transmissable Spongiform Encephalopathies (holes in brain like a sponge)
