LEC36: Transport into Mitochondria, Peroxisomes, & the Nucleus Flashcards Preview

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Flashcards in LEC36: Transport into Mitochondria, Peroxisomes, & the Nucleus Deck (62):
1

what % of proteins go through the secretory pathway versus stay in the cytosol and are targeted elsewhere? where else might they be targeted? 

30% of proteins are handled by secretory pathway at the ER 

70% stay in cytosol or are targeted to mitochondria, nucleus, or peroxisomes 

2

what % of liver volume do mitochondria comprise?

25% 

3

size of mitochondria?

0.5-1 micron diameter

4

function of mitochondria?

generate ATP to be used as cellular source of energy

5

important structural features of mitochondria?

outer mitochondrial membrane, inner mitochondrial membrane with cristae forming big folds, membrane space in between the 2 membranes, mitochondrial matrix in middle of it all

 

A image thumb
6

structure(s) of outer mitochondrial membrane?

porous to molecules up to 5-10 kDa

contains a protein translocation apparatus, TOM 

 

7

structure(s) of inner mitochondrial membrane?

inner membrane is 70% protein, is impermeable even to protons 

inner membrane is highly invaginated into cristae to increase surface area (cristae are variable)

has TIM complex for transport of peptides across it

membrane potential across inner membrane 

8

what is within the inner membrane, re: proteins?

protein complexes of the electron transport chain and ATP synthase complex that catalyzes formation of ATP from ADP 

9

what creates membrane potential across inner membrane?

delta psi 

matrix (inner) is negatively charged; inter membrane space (out) is highly positively charged 

protons moving into the intermembrane space creates membrane potential 

thus inner membrane is sealed tight to protons 

makes it hard to get proteins into the mitochodria 

10

what's different between mitochondria in the skeletal muscle vs. liver?

muscles have lots of cristae; liver do not have much cristae 

b/c liver is more involved in metabolism, and muscle is dedicated to ATP production 

11

what to mitochondrion do in sperm?

in flagella - so sperm can swim

12

what is lodged w/in the inner mitochondria membrane in terms of proteins and molecules?

channel proteins for translocation of metabolites:

pyruvate, malate, acyl-CoA, amino acids 

ions 

ADP/ATP transporter 

 

13

what is in the intermembrane space? 

between the 2 membranes 

contains enzymes that phosphorylate other nucleotides apart from ADP, eg nucleoside diphosphate kinase, which converts GDP -> GTP

14

what is in the mitochondrial matrix?

1) hundreds of enzymes, including those for oxidation of pyruvate, fatty acids, ketone bodies to acetyl-CoA 

2) enzymes that catalyze amino acid oxidation 

3) enzymes of tricarboxyic acid cycle and urea cycle 

4) mitochondrial genome, ribosomes, tRNAs, molecular chaperones for folding newly synthesized & newly imported proteins

15

what is the structure of the mitochondrial genome?

circular 

16,589 bp 

encodes 13 proteins

2-50 copies of genome per mitochondria, so up to 1000/cell

16

what does mitocondrial genome encode?

tRNAs and rRNAs for its own ribosomes

mitochondrial genes are txn into mRNA and tln in matrix on mitochondrial ribosomes

17

what proteins does the mitochondrial genome encode?

subunits for several components of respiratory chain, including cytochrome c oxidase, NADH dehydrogenase, apocytochrome b 

genes for protein synthesis within matrix: 22 tRNAs, 12S and 16S rRNA for mitochondrial ribosomes

18

can gene migration occur btwn the universal and mitochondrial genomes?

no! 

b/c mitochondrial genome has different genetic code than universal genetic code 

i.e. UGA is universal STOP codon; it is Tryptophan in mitochondria 

AGG is R in universal code, STOP in mitochondrial genome

19

how do mitochondria come to exist?

they aren't assembled; divide into 2 from 1 existing mitochondria

20

how are mitochondria inherited in mammals?

maternally

21

what is evidence of mitochondria's prokaryotic origin?

endosymbiotic origin from prokaryotes, like chloroplasts in plants 

evidence: prokaryotic character of irbosomes, molecular chaperones, and circular genome 

22

where do most mitochondrial proteins come from?

synthesized on cytosolic ribosomes before targeting to outer membraen and post-translational import 

23

what is the nature of mitochondrial targeting sequence?

15-35 residuce N-terminal sequence of positive, basic amino acids 

it is cleaved in matrix by an endoprotease 

also has a non-cleaved internal targeting sequence

24

when do proteins destined for mitochondria get transported?

what is the state of the peptide?

post-translationally 

unfolded state 

25

what brings peptide to mitochondria? what does it do?

Hsp70 chaperone 

important b/c protein is imported in unfolded conformation, which Hsp70 maintains 

26

describe how peptide enters mitochondria once it is chaperoned there from the ribosome 

1) Hsp-70 bound peptide binds to receptor on outer mitochondrial membrane; is presented to TOM complex; enter through aqueous TOM40 translocation channel 

no energy involved 

2) positively charged pre-sequence of peptide opens TIM complex on inner mitochondrial membrane

electrophoretic effect occurs, b/c targeting sequence is positive compared to inner part of matrix

TIM opens enough for peptide to get in through inner mitochondrial membrane 

3) once in matrix, mitochondrial version of Hsp70 binds protein 

4) Hsp70 pulls protein into matrix, helps fold it

27

what are all of Hsp70's functions?

1) keep protein unfolded in cytosol 

2) pull protein in to mitochondrial matrix after presequence has entered through TIM complex of inner mitochondrial membrane 

3) helps protein fold after it has entered matrix

28

when does the mitochondrial targeting sequence get cleaved?

once peptide is within the matrix

29

what happens if have a mutation in mitochondrial DNA?

may have genotypic mutation, but it may not express disease phenotype (although it may!) 

penetrance determines this - more copies of a mutation you have in genotype, more experiences in phenotype

30

what is leber's hereditary optic neuropathy (LHON) caused by?

missense mutation in subunit 4 of NADH-coQ reductase 

causes sudden onset blindness, hearing loss, mental retardation 

31

what causes dystonia?

NADH-coQ reductase missense mutation in subunit 4

same mutation as LHON 

32

what does aging due to mitochondria DNA?

rearranges mitochondrial DNA 

probably result of accumulation of reactive oxygen species with time 

humans over age 40 have increased mitochondrial DNA rearrangements compared to humans under age 40 

leads to decreased energy production w/ age 

33

what is the structure of peroxisomes?

small, single membrane organelle 

 

A image thumb
34

what do peroxisomes do?

fatty acid B-oxidation aka they oxidize long chain fatty acids 

this generates hydrogen peroxide which catalase reduces to water and oxygen 

do plasmalogen synthesis - phospholipids found in omst eukaryotic cells, myelin membranes

 

35

what encodes peroxisomal proteins? where are they synthesized? when are they transported? how? 

nuclear genes 

synthesized on cytosolic ribosomes 

imported post-translationally in folded state into peroxisome 

2 C-terminal target sequences: PTS1, it is SKL a.as 

 

36

what does catalse do?

reduces hydrogen peroxide to water and molecular oxygen 

works in peroxisomes 

 

A image thumb
37

what are diseases related to peroxisomes? what are they caused by?

1) zellwegers syndrome - no import of any peroxisomal enzyme 

2) adrenoleukodystrophy (ALD) - oxidation of very long chain fatty acids is defective; ALD gene is membrane transporter for long-chain fatty acyl CoA synthase from cytosol to peroxisome matrix

38

what does nucleus separate?

genetic material inside, separate from cytosol 

separates txn and tln, allowing another level of regulation compared w/ prokaryotes 

39

what activites occur inside nucleus?

transcription

mRNA splicing 

DNA replication 

ribosome biosynthesis

40

when does nucleus form?

nucleus is disassembled prior to mitosis, reforms afterward

41

what is size of nucleus?

largest organelle in cell 

5-10 um diameter

42

what is membrane of nucleus like?

double lipied bilayer

inner = nuclear lamina, gives nucleus tensile strength

outer layer = rough ER membrane, they are contiguous

43

what/where is nuclear lamina?

lines inner surface of nuclear membrane 

meshwork of intermediate filament type proteins, lamins 

 

44

how does transport into/out of nucleus occur?

via aqueous pores, nuclear pore complexes 

45

what is the organization of chromatin?

nucleus houses all all genes, in 46 chromosomes

each chromosome is very long and thin; presents packing problem

DNA is packaged into chromatin which is further compacted into a 30 nm fiber

these 30 nm fiber are interphase nucleus chromatin structure 

46

what is the nucleolus? 

where rRNA is transcribed, synthesized

where ribosomal subunits are assembled

most prominent strucutre within nucleus

47

describe the chormatin of the interphase nucleus

euchromatin (10% of which is transcribed) and heterochromatin, which is highly condensed and not trnscribed 

highly condensed chromatin is in close apposition to the lamina 

euchromatin and heterochromatin define the interphase nucleus 

during mitosis, chromosomes condense into metaphse chromosomes that're more highly organized

48

where are the genes that encode rRNA?

in nucleolus, 200 copies of these  genes, including 5.8S, 18S, 28S on 5 different chromosomes 

all are synthesized as a 45S precursor 

these locii localize to the nucleolus 

49

describe process of ribosomal RNA biogenesis 

they congregate in the nucleolus of the nucleus 

ribosomal mRNA are synthesized on chromsomes all around the nucleus by RNA Pol II 

the mRNA are exported from nucleus; translated on cytplasmic ribosomes 

ribosomal proteins then reenter nucleus, are transported to nucleolus, as does 5S RNA 

there, assemble w/ rRNA 

individual 40S and 60S pre-ribosomal aprticles are then delivered to cytosol 

 

50

how many protein molecules enter nucleus / minute?

how many mRNA? 

tRNA?

how?

60,000 protein 

50-250 mRNA 

1000 tRNA

all via 3,000 nuclear pore complexes

51

what is the nuclear localization sequence?

can exist anywhere on protein 

usually 4-8 a.as, rich in Arg and Lys, usually contains Pro

 

52

in what conformation do proteins enter nucleus?

folded

53

are nuclear export signals = NLS?

no

nuclear export signal is distinct from nuclear localization sequence 

54

when/is the nuclear localization cleaved?

never! 

because proteins shuttle between nucleus & cytoplasm or have to reenter nucleus as it reforms after mitosis

55

how do proteins enter/exit nucleus?

through nuclear pore complexes: 8 subunits long ring 

bidirectional transport occurs through NPCs

 

56

what is structure of the NPC?

nuclear pore complex

3000-4000 per nucleus 

each composed of ~100 diff proteins, collective mass is 125 Md

has an 8 fold symmetry, contains central aqueous channel of 9 nm diameter and 15 nm length 

is several ring assemblies that occupy cytoplasmic face, inner core and nucleoplasmic face of structure 

filamens radiate out from both cytoplasmic and nucleoplasmic sides

57

describe process of protein entry into the nucleus via the NPC

if protein is made in cytosol, wants to go back to nucleus to ribosome 

1) protein binds to karyopherin receptor 

karyopherin enters through NPC w/ protein attached 

2) RAN-GTP causes disassembly of karyopherin complex, cargo protein is released

3) karyopherin is bound to RAN-GTP, they undergo export through NPC back to cytosol 

4) cytosolic RAN-GAP protein promotes disassembly of karyopherin:RAN complex by catalyzing GTP hydrolysis in Ran

karyopherin now free to recycle, pick up new cargo; Ran GDP can return into nucleus,  and Ran ntd exchange factor, RAN Nef, catalyzes formation of Ran GTP to start cycle again

58

what is Ran? what are its forms/where? relations to GAP and GEF?

a small GTPase w/ co-factors that regulate nucleotide hydrolysis (Ran GAP) and ntd exchange (Ran GEF) 

Ran GAP: cytosol, so Ran GDP

Ran GEF: nuclear, so Ran GTP

 

59

during process of secretion: 

is there a signal seuqnce? is signal sequence cleavable? co or post translational?

yes, signal sequence

yes, cleavable, 

co-translational

60

during transport into mitochondria, 

is signal sequence cleavable? 

co or post translational?

yes cleavable 

post translatoinal

61

re: transport to nucleus, 

is signal sequence cleavable? co or post translational?

not cleavable 

post translational

62

for peroxisome, 

is signal sequence cleavable? 

co or porst translational?

not cleavable, and at C terminus

post translational

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