Proteins are imported into organelles by what 3 mechanisms?
1. Transport via nuclear pores - selective gates which actively transport proteins in both directions
2. Transport via protein translocators - protein translocators within organelle membrane directly transport proteins from cytosol into organelle. Occurs co-transitionally for the ER and post-translationally for the mitochondira and peroxisomes.
3. Transport via vesicles - vesicles containg proteins from lumen and membrane of one organelle pinch off and then fuse with the membrane of a second orgnaelle (ex. ER to Golgi, golgi to plasma membrane)
How does a cell know where to deliver each newly synthesized protein?
Signal sequences/sorting signals - used to direct proteins to correct organelle, 3-60 amino acid stretch in protein, may be removed by a signal peptidase
Describe mitochondria's structure and function.
Function: Power plants of the cell - 90% of the energy
Structure: Outer membrane - separates mitochondrian from cytosol, has porins (channels for small molecules)
Inner membrane - encloses matrix space, convoluted (cristae), contains proteins for e- transport chain and ATP synthase, transport proteins, electrochemical gradient maintained across it
Intermembrane space - small space located between inner and outer membranes, contains cytochrome C, component of e- transport chain
Matrix space - high concentration of enzymes including those that break down fatty acids and those in the citric acid cycle, location of mitochondrial DNA genome
Briefly describe the conversion of fatty acids and pyruvate to ATP
Describe the role of mitochondria in Apoptosis
The release of cytochrome C from the intermembrane space into the cytosol triggers activation of the caspase cascade, an intracellular proteolytic cascade responsible for cleaving key cellular proteins
Describe the transport of proteins into mitochondria
Proteins destined for mitochondria are typically synthesized with an N-terminal signal sequence and transported post-translationally. Transport is mediated by two protein translocators - TOM and TIM.
Precursor protein binds to a receptor associated with TOM (transporter in outer membrane). TOM/receptor complex diffuse laterally until it encounters a TIM (transporter in inner mem).
Precursor protein is translocated across both membranes, signal sequence first - chaperone proteins help pull the protein. Signal sequence is cleaved off by mitochondiral signal peptidase and protein folds into final confirmation.
Describe the mitochondrial genome
Very small circular double stranded DNA
Encodes 2 rRNAs, 22 tRNAs, 13 mRNAs
Little regulatory sequence
Genetic code is slightly different (tRNAs don't carry same proteins)
~10-20 copies per mitochondrion
Makes some of the components for the e- transport chain
Describe the replication of Human Mitochondrial DNA
Mitochondria carry out their own DNA rep, transcrip, and protein synth in the matric using mitochondria-specific proteins encoded by the nuclear genome.
Replicaiton occurs throughout cell cycle, look to double # of mitochondria by mitosis, not tightly controlled.
Origin of replication on each strand, mtDNAs chosen at random for replication
Describe the transcription of mtDNA
Both strands transcribed from single promoter on each
Produces 2 giant RNAs, each a transcript of one DNA strand
RNAs cleaved into rRNAs, tRNAs, and mRNAs
Describe the translation of mtDNA
Occurs in matrix
Uses tRNAs and rRNAs encoded in mtDNA
Produces 13 polypeptides - all are subunits of complexes involved in oxidative phosphorylation >>> targeted to inner membrane
What is the function of peroxisomes?
Oxidative degradation - break down a variety of molecules (toxins, alcohol, fatty acids)
Production of H2O2 (RH2 + O2 >>> R + H2O2)
Catalast driven rxns (H2O2 + RH2 >> R + 2 H2O) or (2 H2O2 >> 2 H2O + O2)
Beta oxidation: very long chain fatty acids > acetyl CoA
Synthesis of cholesterol, bile acids, and lipids: ex, plasmalogen for myelin sheaths
How are proteins imported into peroxisomes?
Post-translationally via specific protein translators
Describe disorders of peroxisome biogenesis
Dramatic loss of peroxisome function due to defects in proteins required for peroxisomal biogenesis - peroxisomes lack many peroxisomal enzymes or absent from cells
Ex. Zellweger syndrome - perox enxymes synthesixed normally but not imported, empty peroxs, lethal in early infancy
Describe deficiency of single peroxisomal proteins...
Less severe phenotype, defect in synth, import, or fxn of one perox protein, partially fxnal peroxs
Ex. X-linked adrenoleukodystrophy (ALD)
perox lack membrane protein involved in degradation of very long chain fatty acids
build up of long chain fatty acids
Leads to demyelination of neurons, dysfunction of NS, adrenal insufficienct
lethal in mid childhood
What are ways to treat X-linked adrenoleukodystrophy?
1. Allogeneic hematopoietic stem cell transplant (HCT) - high morbidity due to complications, compatible donor cells not always available, must be performed at early stage due to brain lesions
2. Gene therapy - Stem cells collected from pt, corrected ex-vivo w/ HIV derived lentivirus, bone marrow eradicated, and own stem cells put back
First successful clinical test of HIV-derived vector