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63 Cards in this Set
- Front
- Back
Secretion process |
-Synthesis of proteins on ER and co-translational insertion -Production of vesicles (formation of golgi cisterni) -Golgi enzymes move countercurrent fashion |
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Factors that allow proteins to interact with membrane receptors |
shape, size, charge |
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Destination of post-golgi vesicles |
1. Plasma membrane: exocytosis and fusion (become integral membrane proteins) 2. Endosomes: formation of lysosomes
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v-snares and t-snares |
control fusion of membranes along with other docking proteins |
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Discovery of lysosomes |
Isolated from livers, hypotonic lysing increases enzymatic activity of protein phosphatases --> special population of vesicles associated with lytic activity |
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Protein phosphatase |
present in lysosomes and trans-golgi cisterni |
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Primary and secondary lysosomes |
Termed by electron microscopists Process later examined dynamically and determined to be continuous maturation Shift terms to pre-lysosome and lysosome |
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Determining destination of golgi-derived vesicle |
proteins within vesicle, receptors and proteins on vesicle membrane |
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Hurler syndrome |
individual lacks capacity to make specific lysomal enzymes - lysosomes are swollen due to accumulated substrate |
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Autophagy |
alternative to using lysosomes to degrade substrate Wrapping substrate with ER and expelling |
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Correcting hurler syndrome |
add missing enzyme to external space - it will reach its destination through endocytosis can't really be used in vivo |
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I cell deficiency |
single gene trait, lysosomal enzymes are all secreted --> swollen lysosomes
tx: addition of WT enzyme with mannose-6-phosphate tag to extracellular space |
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Mannose-6-phosphate |
acts as competitive inhibitor of endocytosis if terminal phosphate is removed - protein is secreted
raise MUT in presence: restore normal function |
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Cis-golgi enzyme |
adds phosphorylated n-acetylglucosamine to mannose present on lysosomal proteins through esterification reaction |
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Trans-golgi enzyme |
cleaves n-acetylglucosamine from phosphorylated mannose leaving mannose-6-phosphate tag on lysosomal protein |
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M6P receptors |
bind vesicle proteins - move to endosome -dissociate during acidification and recycled to golgi |
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M6P receptor recycling |
driven by retromers, site= trans golgi |
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M6P tag |
marks vesicles as non-secreted cleaved in functioning lysosome preventing return to golgi |
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Post-translational import |
process of putting proteins into different compartments like mitochondria, chloroplast, peroxisomes and nucleus
Proteins synthesized in cytoplasm and incorporated as intact proteins |
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Production of mitochondria/chloroplast proteins |
mainly occurs in cytoplasm (like 95-8%) transfer proteins are responsible for targeting |
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Co-translational import proteins |
not membrane associated, fold up after synthesis, no glycosylation |
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HSP70 |
example of carrier protein that bring cytoplasmic transcripts to target organelle
facilitates insertion of protein into mitochondrial matrix Unfolds proteins on cytoplasmic side |
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Simple version of post-translational import |
Protein binds to organelle, unfolds, threaded through membranes and refolded within appropriate compartment |
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Target determination of post-translational import protein |
Determined by sequence of transcript and presence of specific membrane proteins that allow insertion to occur |
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n-terminus |
responsible for insertion penetrates membrane first through translocon |
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comparison of outer membrane protein and inner membrane protein of mitochondria |
Differ in length of necessary sequence for targeting Indicates that specific sequence does not encode target - sequence encodes SHAPE which determines target |
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Common things to targeting motifs |
alpha-helical, positively charged, polarized hydrophobicity |
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Cytochrome C |
ignores typical post-translational import pathway CC1 typically inserted into mitochondria inner membrane
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Transit peptides in Cytochrome C |
Start transit peptide at n-terminus and immediately downstream is stop transit peptide |
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Removal of CC1 stop transit peptide causes |
CC1 to be localized within the inner matrix |
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Multiple sites of mitochondria protein insertion |
outer membrane, intermembrane space, inner membrane, matrix (default) |
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Multiple sites of chloroplast protein insertion |
outer/inner membranes, intermembrane space, stroma membrane, thylakoid, stoma lamellae |
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Start and stop transit peptides |
Determine site of insertion of post-translational import protein |
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Post-translational import motifs are called... |
transit peptides |
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Co-translational import motifs are called... |
transport peptides |
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Endopeptidase |
removes transit peptides after insertion preventing movement out of the organelle (cleavage causes change in conformation) |
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TOM TIM |
Translocon of outer and inner mitochondrial membranes respectively align to allow protein to be inserted though both membranes at same time |
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Why do TOM and TIM align? |
Prevent proteins from refolding before reaching matrix space |
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TOC and TIC |
chloroplast counterparts of TOM and TIM, also align to allow proteins to pass into stromal space |
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Inner membrane (of mitochondria) |
site of E generation using ATP synthase proteins and proton gradient insertion of translocon pores does somewhat dissipate proton gradient |
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Porins |
allow water to move through out membrane, inserted via TOM prevented from travelling through inner membrane by chaperones in inner membrane space |
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Mitochondria proteins that are not targeted to the matrix |
Either prevented from traveling through TIM (chaperones encourage protein folding) or are pulled into the matrix only to be pushed out again using stop transit peptide |
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Treatment of mitochondria with ionophores |
Ionophores (DNP, CCCP) block import if protein hasn't yet started insertion Block necessary energy requirements? |
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Change conformation of proteins in matrix |
endopeptidase sometimes allows for insertion into inner membrane |
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ATP synthase activity |
required for continued insertion of proteins into mitochondria |
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Dihydrofoliatereductrase |
cytoplasmic enzyme that creates color when exposed to a second enzyme attach to start transit peptide: expressed in matrix
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Methotrexate |
binds to dihydrofoliatereductrase preventing functionality and unfolding - can't observe color in mitochonria
used to prove that protein unfolding is necessary for post-translation import |
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New n-terminus |
generated after cleavage of start transit peptide hydrophobic: insert into inner membrane or other points in cell/mitochondria |
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Chloroplast post-translational import problems |
Not all membrane pairs have a site of energy production (like inner membrane of mitochondria) Require different mechanisms for import |
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Insertion into thylakoid |
secondary event mediated by SEC proteins, SRP and proton gradient driven pathway |
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Peroxisomes |
Involved in oxidative rx (burn up toxic substances) Major site of oxygen consumptions |
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Disproven theories of peroxisomes |
must be generated from old peroxisomes proteins arrive exclusively through post-translational import |
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Peroxisome membrane proteins v. inner peroxisome proteins |
membrane: co-translation import but not through golgi inner: post-translation import |
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Products of peroxisome reactions |
Peroxide, phenols ethanol to acid aldehyde fatty acids to acetyl CoA |
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Plant use of peroxisome |
breakdown of oils in seeds |
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PEX proteins |
important in getting proteins into peroxisome compose enzymes w/in peroxisome, channels proteins, chaperone proteins |
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Peroxisome formation |
Fission: grow and divide (actin cables pull one into budding cell) |
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Defective PEX |
Defects in PEX3, PEX 6 and PEX 19 --> no peroxisomes |
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Cells without peroxisomes |
perform transvection with restorative proteins and peroxisomes are recovered
new peroxisomes can be generated from p-domain of smooth ER |
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p-domain of Smooth ER |
PMP rich, bleb off as pre-peroxisomes and fuse together using snare proteins |
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PMP proteins |
rich in p-domain of smooth ER involved in peroxisome function and formation |
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Dynamin and peroxisomes |
would form loop around budding section and cuts off new peroxisome from original (in fission)
DNM1 and VPS1 used instead |
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DNM1 and VSP1 |
used by peroxisomes instead of dynamin
DMN1 used for same function in mitochondria, similar alpha-helical structure to dynamin |