MBB11003 -Molecular and Cell Biology Flashcards

Question

What did Avery, MacLeod and McCarthy investigate?

Answer 1

what was causing the transformation Griffith had seen aka what the transforming principle actually was -they concluded it was DNA

Answer 2

which component of a bacteriophage is injected into a bacteria -using bacteriophage T2 (host: E.coli) -they radiolabelled its DNA or protein, infected bacteria, separated phage ghosts and tested phage ghosts for radiolabelled DNA/protein -concluded it was DNA

Answer 3

Chargaff's experiment into the proportions of diff bases using paper chromatography -same proportions of purines and pyrimidines -same proportions of A and T, and C and G (diff to eachother though) X-ray crystallography -X pattern (=helix) -regular pattern (=repeated, even structure) -distance between spots (=distance between turns of helix, 3.5nm)

Answer 4

-A-T and C-G hydrogen bonded base pairs (2 H-bonds for A-T, 3 for C-G) -purines and pyrimidines are base paired to eachother -DNA strands run antiparallel to eachother (one 5' to 3', the other 3' to 5') -double helical structure (each phosphate backbone forms a helix) -double helix has major and minor grooves -one complete turn of the double helix is 10.5 base pairs

Answer 5

-centromere (region which links chromosomes to spindle) -telomere (repetitive sequences of DNA at end of chromosomes)

Answer 6

specialised chromosomal region where chromosomes link to spindle microtubules -directs equal segregation of chromosomes during mitosis and meiosis -not necessarily at centre of chromosome

Answer 7

repetitive DNA sequences at the end of linear chromosomes -protect ends of chromosome

Answer 8

-single circular chromosome w/a few million bps (more compact genome than eukaryotes) -plasmids (small circular molecule w/thousands of bps which can be passed to other prokaryotes via conjugation)

Answer 9

-regulate gene expression (eg. transcriptional regulators like lac operon) -cut DNA at specific sequences (eg. restriction endonucleases) -protect DNA (eg. histones)

Answer 10

-grew bacteria in medium containing N15 (which makes DNA heavy) -transfer bacteria to a medium containing N14 (which makes new DNA lighter) -separate heavy and light molecules by ultracentrifugation -look at DNA using UV light -heavier DNA would form bands at bottom, lighter DNA would form bands at top -results showed DNA replication is semi-conservative

Answer 11

add DNA nucleotides one at a time onto bases from 5' to 3' using template strand

Answer 12

generates a primer (made of RNA)

Answer 13

joins new sections of new DNA together -joins "gaps" in sugar-phosphate backbone to make it a continuous molecule

Answer 14

breaks hydrogen bonds between bases to separate the DNA strands -to unwind helix

Answer 15

binds to strands (separated by helicase) to stop strands reannealing

Answer 16

relieves pressure around the replication bubble by making breaks in the DNA molecule and then resealing it

Answer 17

the strand of DNA where 5' to 3' synthesis points towards the replication fork so it can be continuously replicated

Answer 18

the strand of DNA where 5' to 3' synthesis points away from the replication fork so it can not be continuously replicated -must be primed multiple times -forms Okazaki fragments which need to be joined together

Answer 19

replication is continuous on leading strand but discontinuous on lagging strand (-opposite on other side of replication bubble)

Answer 20

each time the primer is removed (because primer is RNA so must be removed), a gap is left at the end of the chromosome which is impossible to fill (due to DNAP only being able to work from 5' to 3') so a small piece of DNA is lost each time HOWEVER only telomeres (repeating DNA sequences at end of chromosomes) are lost which can be extended by telomerase

Answer 21

replenishes the telomeres (which have been lost when the primer was removed) from an RNA template

Answer 22

specialised proteins which form a protective cap on telomeres -this hides telomeres from being detected as "cell damage" (differentiates it from DNA breakages) so that nucleases don't break it down -this recruits telomerase

Answer 23

canonical (Watson-Crick bps -U/A and C/G) non-canonical (wobble bps)

Answer 24

Watson-Crick base paring U with T and C with G

Answer 25

Wobble base pairing due to folding and intramolecular interactions causing unique 3D structures Non-Watson-Crick pairs eg. U with G Eg. long range tertirary structure interactions like the A-minor motif

Answer 26

Common tertiary interaction in RNA where two consecutive adenine residues interact with adjacent base pairs in minor-groove (elsewhere in the RNA molecule) through ionic interactions -an example of non-canonical base pairing

Answer 27

-RNA is synthesised by DNA-dependent RNA polymerase (which has an active site containing a short RNA/DNA heteroduplex) -genetic sequence in sense strand is transcribed to RNA by nucleotide triphosphates (NTPs) being selected by base pairing with the template strand (transcription bubble) and being added to 3’ end of RNA strand -RNAP is targeted to promotor region and when it reaches the terminator region, it is released (means transcription is not across whole genome but only the section needed to make the protein)

Answer 28

5 subunits: -2 alpha (which bind transcription factors) -beta and beta’ (catalytic) -1 omega (responsible for assembly and stability)

Answer 29

to bind transcription factors

Answer 30

catalytic roles

Answer 31

assembly and stability

Answer 32

targets RNAP to gene promotors, which enables binding IN PROKARYOTES -causes RNAP to have an open, active conformation -DNA in active site forms transcription bubble and short RNA primer is formed -sigma factor is releases, so RNAP moves away from promotor (promotor clearance)

Answer 33

3 nuclear RNAPs (plus others in plants) (have different RNAPs in mitochondria)

Answer 34

RNA polymerase 1 -transcribes rRNA RNA polymerase 2 -transcribes mRNA and non-coding RNAs RNA polymerase 3 -transcribes tRNA and 5s rRNA

Answer 35

-they transcribe different RNAs -each have common and unique subunits

Answer 36

enables RNAP to bind IN EUKARYOTES for RNAP 2… -TATA box binding protein (TBD) in TF2D binds to TATA box in DNA, causing DNA to bend, which allows other factors to be recruited -preinitiation complex (PIC) assembles at promotor using RNAP2 and gTFs

Answer 37

A/T rich region within promotor region of eukaryotic gene promotors -which TATA box binding protein (TBD) in TF2D binds to

Answer 38

-activators (bind to enhancers in DNA sequence to attract RNAP 2 to DNA) -mediators (allows activators, gTPs and RNAP 2 to communicate) -general transcription factors -chromatin-modifying proteins

Answer 39

COUPLED: in prokaryotes mRNA is translated into a protein whilst it is being made by RNAP -causing rapid gene expression -quick responses to changes in environment COMPARTMENTALISED: in eukaryotes mRNA is transcribed in nucleus and then translated in cytoplasm (separate as organelles are compartmentalised) -allows regulation at diff steps -less direct but more diverse

Answer 40

-5' end is capped -introns are removed (pre-mRNA splicing) -3' end is processed (cleavage and polyadenylation)

Answer 41

helps mRNA to be distinguished from other types of RNA in the cell -this helps it to be further processed and exported and aids its translation

Answer 42

-phosphate is removed from 5' end by a phosphatase -GMP (guanosine monophosphate guanine) is added (ie. guanine added) in reverse linkage (5' to 5' insread of 5' to 3') by a guanyl transferase -methyl group is added to guanosine by a methyl transferase (usually at position 7) ∴ known as m7G cap

Answer 43

extra guanine nucleotide added to 5' ∴G cap nucleotide is methylated usually at position 7 ∴m7

Answer 44

a mRNA molecule only codes for one polypeptide (in prokaryotes, one mRNA can be translated into lots of different proteins)

Answer 45

large complexes made up of RNA and a spliceosome (protein) which carry out pre-mRNA splicing by removing introns (keeping exons) -smaller RNA/protein complexes (snurps) assemble/disassemble active spliceosomes

Answer 46

small nuclear ribonucleoproteins aka small RNA/protein complexes -assemble/disassemble active spliceosomes

Answer 47

splice site sequences allow recognition of intronsic and exonic sequences -intron contains a 5' splice site, a branch point and a 3' splice site -the 5' and 3' splice sites are highly conserved and allow this recognition

Answer 48

2 steps (transesterification rxn) -2' hydroxy group of branchpoint adenosine attacks 3' phosphate of exon -this 5'-2' phosphodiester bond gives a looped lariat -the 3' hydroxy group (generated from the first step) attacks the 5' phosphate of the 3' exon and the lariat is released (lariat is then degraded by enzymes)

Answer 49

rRNA (LARGE -25s and 18s appear on agrose gel; 5.8s and 5s appear on acrylamide gel) tRNA (SMALL -only appears on acrylamide gel) N/B: mRNA is not clearly visible -cells transcribe lots of diff mRNAs which vary in length and are very unstable

Answer 50

-in vitro translation (cell extract isolated and its mRNA is degraded, synthetic C14-labelled RNA/aas are added and then protein is precipitated and collected) -ribosome binding (isolated purified ribosomes are incubated with radiolabelled amino-acyl tRNAs and mRNA codon, forming a stable ribsome/aa-tRNA codon-complex which can be separated to check its radioactivity)

Answer 51

UAA UAG UGA

Answer 52

61 -there are 64 possible codons but 3 of them (UAA, UAG and UGA) code for the stop codon -degenerate: multiple codons code for same aa (synonymous codons tend to vary on 3rd position of codon) -some codons just code for one aa (eg. AUG only codes for Met)

Answer 53

-cloverleaf secondary structure (5' and 3' drawn together, aa s attached to 3' OH of 3' terminal adenine) -specific nucleotides are post-transcriptionally modified (allows wobble in base pairing) -folded into L-shape -caused by coaxial stacking of helices and base pairing between ends of loops

Answer 54

aminoacyl-tRNA synthases charge tRNAs by adding on an amino acid -requires ATP (which is converted into ADP and Pi) -aa is linked to tRNA by an ester linkage between carboxy group of aa and 3' hydroxy group of terminal nucleotide of tRNA

Answer 55

for gene expression -allows aa to be delivered to ribosome for translation to produce polypeptide chain

Answer 56

-2 RNP (ribonucleoprotein) subunits: large subunit (where peptide bond formation occurs) and small subunit (where codon/anticodon binding occurs) -3 non-overlapping tRNA binding sites at subunit intersurface: A (where acyl-RNA binds), P (where peptidyl RNA binds) and E (where non-charged tRNA bind before leaving) -peptidyl transferase centre (PTC) is RNA-rich (RNA catalyses formation of peptide bond) -have a polypeptide tunnel -have a decoding centre

Answer 57

-rRNA transcription and early rRNA processing occurs in nucleoli -processing and assembly occurs in nucleoplasm and cytoplasm -functionally active ribosomes are only generated once they've left the nucleus (translation occurs in cyctoplasm)

Answer 58

-2 tRNAs bind to A and P sites (known as pre-translocation state) (each tRNA is bought to ribosome by EF1A, which hydrolyses 1 GTP) -peptidyl transferase catalyses the formation of a peptide bond between the amino acids attached to these 2 tRNAs -ester linkage between amino acid and tRNA breaks -translocation: ribosome moves along mRNA (requires EF2, so 1 GTP is hydrolysed) so these 2tRNAs enter the P and E sites (post-translocation sites) and the tRNA in the E site leaves -another tRNA binds in the A site and this repeats

Answer 59

A -where acyl-RNA binds P -where peptidyl RNA binds E -where non-charged tRNA bind before leaving (exit site)

Answer 60

elongation factors EF1A (brings aminoacyl-tRNA to ribosome) and EF2 (needed for translocation) -each hydrolyse one GTP

Answer 61

elongation factors EF-Tu and EFG

Answer 62

initiator methionyl-tRNA -recognises AUG codon, targeted to P site elongator methionyl-tRNA -binds to AUG codon, bought to A site

Answer 63

Shine-Dalgarno (SD) sequence in mRNA is recognised by 16S rRNA by base pairing with nucleotides at the 3' end of it -SD seq is typically AGGA

Answer 64

-initiator tRNA (bound to eIF2 and small subunit) is assembled at 5' end of mRNA by interacting with the cap-binding complex (CBC) -preinitiation complex scans along mRNA using CBC's helicase activity until it locates the initiation codon (AUG) in an appropriate context -after the initiation codon is selected, the large subunit of the ribosome is recruited

Answer 65

-termination/release factors (PROTEINS) recognise stop codons -initial binding of release factor (RF1 or RF2 depending on which stop codon) triggers peptide hydrolysis (recognition) -RF3 (a GTPase) allows RF1/RF2 to be released from ribosome -additional factors dissociate ribosome complex

Answer 66

-termination/release factors (PROTEINS) recognise stop codons -initial binding of release factor eRF triggers peptide hydrolysis (recognition) -eRF3 (a GTPase) allows eRF to be released from ribosome -additional factors dissociate ribosome complex

Answer 67

the regulatory sequence in DNA (or RNA transcript) which trans-acting factors interact with

Answer 68

a protein or RNA molecule which interacts with a cis element to regulate the expression of a target gene, different to the gene it was encoded by

Answer 69

in cis -mutations within the same gene in trans -mutations within a different gene

Answer 70

DNA/RNA sequences which affect gene regulation

Answer 71

factors that regulate the expression of a target gene

Answer 72

-trans-acting factors (activators or repressors) can activate or repress translation -substrate-product availability can regulate the expression of enzyme-coding genes (inducers or corepressors up or down regulate expression of enzyme-coding genes)

Answer 73

trans-acting factors which cause the activation of gene expression by interacting with RNAP's alpha subunit to promote DNA binding -promote expression at weak promotors +ve control (transcription increased)

Answer 74

trans-acting factors which cause down regulation of transcription -ve control (transcription decreased)

Answer 75

E.coli gene promotors have bipartite sequences (2 separate nucleotide sequences both recognised by polymerase which act together) -sequences close to eachother are stronger promotors so have high transcriptional activity -sequences divergent to eachother are weaker promotors so have lower transcriptional activity so need to be stimulated by a transcriptional activator

Answer 76

trans-acting factors can activate or repress transcription: -trans-acting activators interact w/alpha subunit of RNAP and promote DNA binding -trans-acting repressors cause down regulation of transcription

Answer 77

-substrates can cause upregulated expression of enzymes (known as inducers) aka increase transcription -substrates can cause downregulated expression of enzymes (known as corepressors) aka decrease transcription

Answer 78

pre-mRNA splicing (introns removed, exons included/excluded) -can occur in diff patterns, resulting in 2 distinct proteins being produced -alternatively, can result in a non-productive pathway, where mRNA is degraded and expression is blocked

Answer 79

bind to specific sequences within pre-mRNA to promote exon inclusion or exclusion -either one or the other of adjacent exons are included

Answer 80

-typically at initiation step -or gene expression can be repressed due to presence of a repressor or small metabolite or diff conditions

Answer 81

-gene product influences its own expression (eg. if too much is produced)

Answer 82

regulatory gene: Pi and lacI Plac lacO structural genes: lac Z, lac Y, lac A

Answer 83

lac Z -codes for beta galactosidase lac Y -codes for lactose permease lac A -codes for acetyltransferase

Answer 84

allolactose (a derivative of lactose generated by β galactosidase)

Answer 85

-is a homoetetramer (has 4 identical subunits associated, but not covalently bound) -has 3 domains: tetramerisation domain (allows protein to be assembled into a tetramer), core domain (where inducer binds) and head domain (where the DNA operator sequence binds)

Answer 86

-DNA binding sites of each subunit are aligned, causing DNA to twist -makes RNAP transcriptionally inactive (it can still bind but isn't active anymore)

Answer 87

-glucose is E.coli's preferred carbon source, so when glucose is present genes required for metabolism of other sugars (eg. lactose) are repressed -when lactose is present as well as glucose, lactose is only metabolised after all the glucose has been used up -causing a diauxic growth curve -lac operon requires absence of glucose and presence of lactose

Answer 88

-adenylate cyclase makes cyclic adenosine monophosphate (cAMP) -cAMP binds to CAP -cAMP/CAP complex binds to CAP site on DNA -this is required for RNAP activity, allowing the structural genes (lac Z, Y and A) to be coded for mutations in the genes adenylate cyclase and CAP block the expression of the lac operon N/B: adenylate cyclase is inhibited in presence of glucose (only occurs when lactose is being metabolised)

Answer 89

-eIF2 binds to Met-tRNA (charged initiator tRNA) and brings it to a small ribosomal subunit -when initiation codon is localised, eIF2 hydrolyses GTP to GDP (eIF2 has a v. low GTPase activity so needs to be stimulates by a GTPase activating protein (GAP)) -this causes a conformational change in eIF2 -this conformational change causes eIF2 and GDP to be released from the initiator tRNA -cells can downregulate translation by depleting levels of eIF2 (known as integrated stress response)

Answer 90

GAP = GTPase activating protein -stimulate activity of GTPases (eg. eIF2)

Answer 91

a guanine exchange factor (GEF) -promotes release of GDP and GTPase is then recharged with GTP

Answer 92

GEF = guanine exchange factor -promotes release of GDP -recharges GTPase with GTP

Answer 93

-protein kinases phosphorylate eIF2 -eIF2-Pi binds tightly to eIF2B (decreasing levels of eiF2B) -this acts as an inhibitor (rather than a substrate) known as an integrated stress response

Answer 94

-to isolate a specific region of DNA (eg. a gene, promotor, intron, etc) -downstream to be able to sequence gene, analyse mutants vs normal genes, express and purify proteins, etc

Answer 95

-DNA is cut using restriction enzymes -DNA is placed into a vector to get recombinant DNA -recombinant DNA is transferred into a host -host w/recomb DNA is selected and replicated

Answer 96

enzymes which recognise a short, specific DNA sequence -4 types (type 2 most commonly used in labs)

Answer 97

restriction enzymes which recognise specific 4-8bp sequence and generate sticky or blunt ends and generate 5’ phosphate and 3’ OH groups -have a polindromic DNA sequence (reads the same 5’ to 3’ on both strands)

Answer 98

a sequence which reads the same 5’ to 3’ on both strands

Answer 99

type 1 and 3 -cleave DNA at random point (far) type 2 -cuts DNA at specific point (close to recognition site) type 4 -cleaves modified DNA

Answer 100

-bind non-specifically to DNA -move along DNA until it finds recognition site -specific binding triggers structural changes in enzyme and DNA -Mg2+ required -5' phosphate and 3' hydroxy ends are generated

Answer 101

to allow vector and insert to stick to eachother

Answer 102

-only hydrogen bonds -no phosphodiester bonds => can be solved by using DNA ligase (eg. phage T4) to reform phosphodiester bonds

Answer 103

-AMP is transferred to a lysine residue in ligase's active site -AMP is transferred to 5' phosphate -AMP-P bond is attacked by 3' OH, forming covalent bond and releasing AMP -ATP is required to replace AMP (∴ ATP is cofactor)

Answer 104

-not enough DNA -DNA mixed w/other molecules -no convenient restriction site -vector self-ligates -incorrect recombinant DNA (wrong orientation)

Answer 105

modifying DNA ends -removing 5' phosphate (only 1 phosphate so no phosphodiester bonds; sticky ends can base pair but can't ligate) -adding 5' phosphate -removing DNA overhang

Answer 106

origin of replication (allows it to replicate inside host) selectable marker (allows cells containing vector to survive) multiple cloning sites (where gene is cloned)

Answer 107

transformation -electroporation (high voltage pulse to induce transient pores in membrane) -chemical transformation (heat shock causes cell membranes to change so DNA can be taken up)

Answer 108

by using a selectable marker on the vector eg. antibiotic resistance genes -can then treat with antibiotic and see which survive (ISSUE: can't distinguish between empty vector and vector containing recombinant DNA)

Answer 109

to amplify DNA in vitro -specific and selective

Answer 110

molecules of DNA double each cycle

Answer 111

-denaturation (double stranded DNA dissociates into single stranded DNA) at 95°C -primer annealing (primers bind to comp seq) at 55-65°C (temp depends on melting temp of primer) -primer extension (DNAP synthesises new strands of DNA from 3' end of primers) at 68-72°C

Answer 112

Taq (used a lot, faster) Pfu (slower but more accurate and better thermostability) -which is used depends on what you need most eg. thermostability, extension rate, etc

Answer 113

-specific to template -at least 17bp long (typically 20bp) -come in pairs (bind opposite strands in opposite directions) -Tm around 60-65°C (primer Tm determines what temp to use for annealing)

Answer 114

primers bind to specific sequence

Answer 115

primers might bind non-specifically to other DNA sequences

Answer 116

primers might not bind at all/not bind efficiently -reduces yield

Answer 117

-blunt-ended cloning is inefficent -blunt-ended cloning is non-directional -PCR products have no 5' phosphate -not an issue if vector has one, but if vector does there is a risk of it self-ligating -taq polymerase adds a 3' alanine overhang to its products (can remove it/some vectors can use it!) to avoid issues: restriction sites can be incorporated into primer

Answer 118

RT-PCR quantitative PCR (qPCR)

Answer 119

-molecular cloning of a protein coding cDNA sequence -analysing mRNA expression

Answer 120

RNA is reverse transcribed into complementary DNA (cDNA) and then PCR is used to amplify specific cDNA sequence

Answer 121

-reverse transcriptase synthesises first strand of cDNA -poly(dT) primers bind to poly(A) tail of mRNA -RNA is removed -second strand of CDNA is synthesised by Klenow fragment of DNAP1 -hairpin formed by reverse transcriptase acts as a primer -ssDNA loop can be digested by a nuclease

Answer 122

amplifcation is exponential but after a while it plateaus because dNTPs and primers run out or DNAP looses activity -qPCR allows you to calculate the relative levels of what you had to start with

Answer 123

using a fluorescent dye (SYBR green, which fluoresces when it binds to DNAs proportionally to amount of DNA -not sequence specific) or fluorescent probes (fluoresces when displaces from template, sequence specific, can multiplex) -fluorescence increases over time

Answer 124

the point at which the fluorescence (from the dye or probes) exceeds the background level -the lower the Ct value, the less cycles are needed to reach the threshold -difference in Ct values between two samples can be used to calculate relative amounts

Answer 125

to make sure it is correct (checking they aren't empty vectors)

Answer 126

a mini prep

Answer 127

-grow lots of bacteria -break bacteria open -genomic DNA (not plasmids -too small and compact) precipitates -get µg of purified plasmid DNA

Answer 128

digest DNA with a restriction enzyme

Answer 129

separate DNA fragments based on their size by using a current

Answer 130

-DNA ladder and samples are loaded into wells -power is turned on and DNA fragments migrate through the gel from the -ve electrode to +ve electrode -fragments are then separated by size (largest fragments towards -ve, smallest towards +ve) -dye is added to visualise

Answer 131

identifies which recombinant DNAs have been successful/which have the gene orientated incorrectly or are just an empty vector by using restriction enzymes -successful recombinant DNA will produce a PCR product whereas an empty vector or incorrect recombinant DNA will not

Answer 132

specialised form of DNA synthesis where the identity of a nucleotide at a specific position can be identified (synthesis can be stopped at a known nucleotide, meaning we know what length the DNA molecule is) (can carry out with A, then repeat with other nucleotides) -use ddNTPs with fluorescent labels -aka Chain Termination Sequencing

Answer 133

deoxynucleotide phosphate

Answer 134

dideoxynucleotide phosphate -a modified version of dNTP where there is not a 3' OH (just H)

Answer 135

-originally 4 separate rxns, each with a diff ddNTP, now 1 rxn with all 4 ddNTPs present, each with a diff fluorescent label -originally radioactively labelled primer -originally T7 DNAP was used, now taq DNAP is used -originally used agarose electrophoresis, now use capillary gel electrophoresis -originally more DNA templates were needed and it was more labour-intensive -now is more specific and sensitive -now is easier, quicker and cheaper

Answer 136

gene expression of DNA/RNA: -PCr-based -hybridisation based gene expression of proteins: -immunological-based -fusion proteins

Answer 137

-Northen Plot Analysis -Microarrays -Fluorescent in situ hybridisation (FISH)

Answer 138

-RNA separated by size using electrophoresis -RNA is transferred to a membrane -allows for blotting -gene-specific probe is added and hybridises to target sequence

Answer 139

-slow -only gives info about 1 gene at a time

Answer 140

-olignonucleotides (short single-stranded DNA/RNA frags) are attached to spot on a chip -each spot has a diff oligonucleotide, specific to a gene -RNA is prepared -fluorescently labelled cDNA is made from RNA -fluorescent cDNA is applied to chip and allowed to hybridise

Answer 141

-not easy to quantify mRNA levels -better for assessing relative levels

Answer 142

a short single-stranded DNA/RNA fragment -used in genetic analysis

Answer 143

-probe is labelled with fluorescent marker and visualised using microscopy

Answer 144

protein-specific antibodies

Answer 145

antibodies which recognise primary antibodies -have a conjugate attached -something we can detect

Answer 146

-proteins are separated by size using electrophoresis -proteins are transferred to a membrane -proteins are detected using a primary (protein-specific) antibody and a labelled secondary antibody (often labelled w/light)

Answer 147

-where protein is (ie. which tissue it's in) -protein levels -about post-translation modification of protein

Answer 148

-secondary antibody is conjugated to a fluorescent marker and is visualised using microscopy -diff fluorophores are used so that multiple molecules can be detected (NOT a live image)

Answer 149

fluorescent chemical compound which re-emits light upon excitation

Answer 150

-fuse 2 protein-coding sequences (CDSs) -protein and fusion protein -use fluorescent fusion proteins eg. GFP

Answer 151

a gene which is easy to visualise or assay which is attached to a regulatory sequence of another gene

Answer 152

protein-protein interactions: -pull down assay -immunoprecipitation -yeast two-hybrid protein-DNA interactions -chromatin immunoprecipitation (ChIp)

Answer 153

-make recombinant DNA -cell lysis done to obtain a cell lysate (fluid containing cell contents) -bind fusion protein (eg. GST) to affinity ligand -wash away unwanted substances to get purified recombinant protein -see what interacts with it

Answer 154

-make recombinant DNA -cell lysis done to obtain a cell lysate (fluid containing cell contents) -bind antibody to affinity ligand -wash away unwanted substances to get purified recombinant protein -see what interacts with it (co-immunoprecipitation)

Answer 155

-using fusion proteins -DNA binding protein is fused to a "bait" and transcription activator is fused to "prey" -if bait and prey interact, transcription of reporter gene occurs

Answer 156

-an antibody or fusion protein is used to purify protein -assay which DNA molecules are associated with that protein by DNA analysis

Answer 157

Western Blot

Answer 158

Quantitative RT-PCR Northern Blot Microassay Luciferase assay

Answer 159

Immunofluorescence Live cell imaging

Answer 160

Immunoprecipitation Pull down assay Yeast-two hybrid ChIp (w/DNA)

Answer 161

ChIp (w/protein)

Answer 162

plasma membrane (cell boundary, controls movement in and out) organelle membrane (compartmentalises cytoplasm)

Answer 163

-act as a barrier -flexible -can self repair -continuous (no edges) -selectively permeable (only certain molecules can pass through)

Answer 164

-lipids -proteins -carbohydrates (linked to lipids as glycolipids or to proteins as glycoproteins)

Answer 165

-rotation -flexion -flip-flop (moving from one leaflet to the other) (RARE)

Answer 166

have hydrophilic, polar head and hydrophobic tail

Answer 167

no. double bonds and no. carbons -more C=Cs = more fluid (unsaturated phospholipids are more mobile) -more acyl chains = more fluid

Answer 168

-phosphatidyl-ethanolamine -phosphatidyl-serine (-ve) -phosphatidyl-choline -sphingomyelin

Answer 169

unsaturated phospholipids have cis-double bonds so are more mobile/fluid saturated phospholipids have longer chains so are less mobile/fluid

Answer 170

-polar head -rigid steroid ring structure -non-polar hydrocarbon chain -smaller than other lipids

Answer 171

makes membrane less permeable -packs between phospholipids, making the surface of the membrane more deformable/rigid (not the whole membrane) -at high temps, stops membrane becoming crystalleine

Answer 172

makes membrane less permeable -packs between phospholipids, making the surface of the membrane more deformable/rigid (not the whole membrane) -at high temps, stops membrane becoming crystalline

Answer 173

micelles -cone-shaped lipids bilayer (more energetically favourable in spheres than planar) -cyclinder-shaped lipids

Answer 174

dynamic movement of proteins in membrane -provides insight into protein's structure (proteins are tagged with GFP and some are bleached -can see the movement of the coloured proteins into the areas of bleached)

Answer 175

proteins inserted directly into membrane by hydrophobic domain -needs strong detergent to remove

Answer 176

proteins associated with integral membrane proteins or covalently bound to lipids inserted in the membrane

Answer 177

clusters of lipids -microdomains form because membrane lipids aren't all the same (aren't homologous) eg. Rafts -formed from cholesterol and spingolipids -more rigid and thicker than rest of membrane, allowing proteins with a long transmembrane protein to be selected

Answer 178

in hypotonic solutions, they form ghost cells which burst, which looses all the haemoglobin so the membranes can easily be looked at by themselves

Answer 179

a cytoskeletal protein which holds cytoskeleton to plasma membrane -in red blood cells, spectrin forms lattice under cell surface, giving them flexibility

Answer 180

-asymmetric -protein always orientated the same way -lipid composition diff in each half of bilayer

Answer 181

a cytoskeletal protein which holds cytoskeleton to plasma membrane -in red blood cells, spectrin forms lattice under cell surface, giving them flexibility

Answer 182

-allows blood groups (O, A, B, AB) to be different to eachother -diff terminal sugars of oligosaccharide chains determines blood groups

Answer 183

phosphatidylserine on platelet membranes provides a site for nucleation for coagulation cascade

Answer 184

aminophospholipids (phosphotidylserine and phosphatidylethanolamine) are transferred to outer leaflet of apoptotic cells' plasma membrane which are detected by receptors on macrophages' plasma membranes

Answer 185

-highly impermeable to ions and solutes (small, uncharged polar) (allows hydrophobic, large, polar substances through)

Answer 186

using liposomes (artificial vesicles)

Answer 187

channel proteins carrier proteins

Answer 188

carrier proteins

Answer 189

combination of membrane potential (voltage across cell) and concentration gradient -established by ionic concentration differences on either side of membrane produced by ion channels, carriers and pumps -involved in lots of processes, such as driving transport, conveying electrical signals (nerves) and making ATP

Answer 190

ions can pass through

Answer 191

ions can pass through easily

Answer 192

ions movement is inhibited

Answer 193

-carriers bind to solute, channels only weakly interact with solute ∴transport is faster with channels -channels passive, carriers passive or active

Answer 194

channels only weekly interact with solute whereas carriers bind to solute

Answer 195

-form hydrophilic pores through membrane -specific for diff ions (∴lots of diff types) -rapid movement down conc/electrochemical gradient -rapidly close and open (intra or extracellular) -stimulated by voltage, ligand binding or mechanisms

Answer 196

voltage-gated channels ligand-gated channels mechanically-gated channels

Answer 197

by undergoing a conformational change

Answer 198

coupled carrier ATP-driven pump light-driven pump

Answer 199

symport -2 solutes transported in same direction antiport -2 solutes transported in diff directions

Answer 200

carriers which only transport one solute

Answer 201

coupled carriers where 2 solutes are transported in the same direction

Answer 202

coupled carriers where 2 solutes are transport in opposite directions to eachother

Answer 203

Na+ gradients

Answer 204

H+ gradients

Answer 205

-largest biomass -occupy lots of diff niches -most variation

Answer 206

separates processes of transcription and translation, enabling alternative splicing

Answer 207

-invagination of membrane around DNA, forming a primitive nucleus (explains double membrane) -one prokaryote engulfing another (endosymbiosis)

Answer 208

using homology hit analysis to compare yeast cells with Archae -further up graph, more similar genes, more commonly related

Answer 209

-DNA is wrapped around nucleosomes (8 subunits of histones) -wrapped twice around each histone -tightly packed -30nm fibre

Answer 210

basic structure of DNA consisting of a segment of DNA being wrapped around 8 histones

Answer 211

dense, tightly packed chromatin

Answer 212

loosely packed, less dense chromatin

Answer 213

dense staining of interphase DNA = heterochromatin less dense staining of interphase DNA = euchromatin very dense staining of RNA = nucleolus (black and white staining)

Answer 214

evidence for heterochromatin and euchromatin -individual chromosomes can be picked out -diff chromosomes occupy specific regions (inherited so region does change) so are found in heterochromatin and euchromatin

Answer 215

location of genes within nucleus changes depending on its transcriptional status

Answer 216

-compartmentalised -dynamic -occupies

Answer 217

sub-nuclear organelles are present in the inter-chromatin space -are dynamic and move in non-random ways using ATP eg. PML bodies, Cojal bodies, etc

Answer 218

by gene tagging and improvements in microscopy

Answer 219

ribosome synthesis

Answer 220

pre-mRNA processing

Answer 221

-not membrane bound -collection of macromolecules incl rRNA genes, precursor rRNA, mature rRNA, RNA processing enzymes, snoRNPs, ribosomal protein subunits, partly assembled ribosomes (sometimes contain mRNA and tRNA)

Answer 222

a non-membrane bound area in the nucleus where ribosomes are produced using ribosomal RNA

Answer 223

a double unit membrane around the nucleus perforated by pores and supported by lamella (fibrous meshwork)

Answer 224

-double unit membrane -has pores -supported by lamella (meshwork of lamin fibres) -lamella is found between the nuclear envelope and heterochromatin and is responsible for the nuclear envelope's asymmetric nature (as diff proteins need to be in inner membrane to associate with lamella)

Answer 225

-ensures nuclear envelope's double membrane is asymmetric (diff proteins present in outer and inner membrane as proteins in inner membrane are near lamella eg. lamella associated proteins aka LAP) -aids structural integrity -involved in gene regulation

Answer 226

mutation in gene encoding lamin A (is an example of a laminopathy)

Answer 227

inherited diseases due to damage to lamin in nucleus (eg. amino acid mutations) -diverse range eg. Hutchinson-Gilford progeria syndrome due to mutation in gene encoding lamin A

Answer 228

SEM, Cryo-EM, Gold Transmission EM

Answer 229

-on cytoplasmic side, have fibrils -on nuclear side, have nuclear basket of fibrils

Answer 230

to control access in and out of the nucleus in a size-dependent manner -particles > 50,000g/mol can't enter nucleus by simple diffusion but can by active signal-dependent transport (which causes nuclear pores to open up to 26nm in diameter) where a specific peptide sequence (nuclear localisation sequence) is required for entry

Answer 231

-phospholipid synthesis adds to cytosolic side of phospholipid bilayer -scramblase catalyses these phospholipids to be flipped in the bilayer, resulting in symmetric growth of both halves of the bilayer (lipids are equilibrated) -new membrane is delivered via exocytosis -flippase catalyses specific phospholipids to be flipped to cytoplasmic monolayer, which ensures asymmetry of bilayer is maintained

Answer 232

flips newly synthesised phospholipids in bilayer to equilibrate lipids, causing symmetric growth of bilayer

Answer 233

flips specific phospholipids to cytoplasmic monolayer, ensuring the membrane's asymmetry is maintained

Answer 234

aminophospholipid translocase (a flippase) transfers a phosphatidylserine from extracellular leaflet of the membrane to its cytosolic leaflet

Answer 235

a deficiency in scramblase -results in increased bleeding

Answer 236

it is continuously breaking and reforming

Answer 237

hollow tubes and flattened sacs -connected to nuclear envelope

Answer 238

-rough (has ribosomes sitting on pores of ER) -smooth

Answer 239

-quality control -protein synthesis -lipid synthesis -storage -detoxification

Answer 240

-phospholipid and cholesterol synthesis -steroid hormone production -glyceride and glycogen synthesis -glyceride and glycogen storage -calcium storage

Answer 241

proteins which aid the folding of newly synthesised amino acid sequences into tertiary and quaternary structures -associate with proteins in lumen of ER if they are not yet folded

Answer 242

export translocate misfolded proteins so that they are degraded -means only correctly folded proteins enter the secretory pathway

Answer 243

-after stimulation, acinar cells release Ca2+ -this causes vesicles to fuse with cell surface membrane -digestive enzymes are released

Answer 244

overload of Ca2+ (can be due to failure of Ca2+ pumps) causing alcohol/biliary disease

Answer 245

-vesicles bud off from donor compartment of endoplasmic reticulum -vesicle moves through cytoskeletan -vesicle's coat is discarded (enables it to fuse) -vesicles fuse to target compartment of Golgi apparatus, releasing the contents of its lumen into the target compartment

Answer 246

lattice-like cage of peripheral membrane proteins (specialised proteins) around the membrane forming the vesicle -coat aids formation of vesicle -coat must be discarded to reveal SNARE before vesicle can fuse with target compartment

Answer 247

-clathrin -COPI -COPII

Answer 248

move substances forwards and backwards through the golgi apparatus and back to the ER

Answer 249

bud from ER

Answer 250

transmembrane proteins (typically with large cytoplasmic domain and small lumenal domain) which ensure the correct vesicle fuses with the correct target component

Answer 251

v-SNAREs (vesicle-SNAREs) t-SNARES (target-SNAREs)

Answer 252

v-SNAREs (in vesicle's membrane) recognises complementary t-SNARE (in target membrane) and they bind, forming a SNARE pair

Answer 253

-endoplasmic reticulum (backwards -retrograde) -cell membrane, etc (forwards -anterograde)

Answer 254

transport from the Golgi apparatus back to the endoplasmic reticulum

Answer 255

transport from the Golgi apparatus to the plasma membrane

Answer 256

composed of flattened sacs called cisternae, which together make a golgi stack* -cis golgi network -cis cisternae* -medial cisternae* -trans cisternae* -trans golgi network

Answer 257

-cis cisternae -medial cisternae -trans cisternae

Answer 258

-modification and packaging of secreted proteins (diff modifications occur in diff, specific regions) -renewal and modification of plasma membrane -delivery of substances to other organelles (especially in the endocytic pathway)

Answer 259

trans face

Answer 260

-signal-mediated diversion to lysosome or to secretory vesicle -constitutive secretory pathway to plasma membrane

Answer 261

-nutrients (macromolecules) -signals (growth factors, morphogens, etc) -antibodies -enzymes -viruses -bacteria -membranes

Answer 262

-is delivered to early endosome (a membrane-bound organelle) -is degraded -is recycled (sent back to cell surface) -is sent to apical domain of plasma membrane (transcytosis)

Answer 263

vesicular transport from one side of a cell to the other -substance is taken up via endocytosis (at basolateral domain of membrane), transported across using vesicles and released at the other face via exocytosis (at apical domain of membrane)

Answer 264

large scale endocytic pathways eg. phagocytosis, macropinocytosis small scale endocytic pathways eg. clathrin-mediated, caveolar-mediated, etc

Answer 265

a form of endocytosis where large vesicles called phagosomes ingest large particles, such as bacteria and apoptotic cells

Answer 266

-pathogen is coated in antibodies (opsonisation) to make it detectable by phagocytes (or other cells) -bind to phagocyte surface receptors, which detect them -phagocyte produces pseudopods (ruffles) which encircle the pathogen (as they encircle it, more and more receptors bind to antibodies on surface of pathogen) until it becomes enclosed by the phagocyte's membrane

Answer 267

-cover slip is opsonised (coated w/antibodies) -phagocyte is attached -phagocyte will try to engulf antibodies but is unable to -we are then able to see what is recruited at this point and what mechanisms are in place

Answer 268

-total SA of cell membrane doesn't change -membrane taken in quicker than it can be synthesised ∴must be recycled

Answer 269

-macrophages taking up beads to mimic phagocytosis -SA of beads is equiv. to membrane taken in -total SA of macrophage stays the sme -membrane is taken in quicker than it can be synthesised so it must be recycled

Answer 270

an actin-driven form of endocytosis where extra-cellular material is taken in non-selectively

Answer 271

-more receptors are made, which are clustered in clathrin-coated pits (precursor to vesicle) -when substance (eg. LDL) binds, coated pits bud off to form clathrin-coated vesicles -vesicles discard their coats before fusing with endosome -in early endosome, substance dissociated from receptors -transport vesicles, containing receptors, bud off from endosome and recycle the receptors, bringing them back to the membrane -substance is released from endosome into lysosomes

Answer 272

-clathrin triskelia (three heavy and three light chains) polymerise into a lattice -adaptor proteins attach the clathrin coat (lattice) to the receptors

Answer 273

-reduction in pH aids maturing of endosomes -early endosome requires lower pH than plasma membrane for substances to dissociate from receptors -lysosomes require even lower pH for optimum enzymatic digestion

Answer 274

help SNARES to mediate fusion -are specific -diff Rabs for diff organelles

Answer 275

-energy production (H+ gradient produced across inner mitochondrial membrane which drives ATPase) -involved in apoptosis

Answer 276

-have a double membrane -inner membrane contains enzymes and transport proteins and is highly folded -folds are called cristae (increase SA) -outer membrane encloses organelle and contains enzymes for mitochondrial lipid synthesis and has porins (which allow entry of molecules <5000kDa) -inter-membrane space -matrix contains mitochondrial DNA, tRNAs, ribosomes (39S and 28S), enzymes and metabolites

Answer 277

-highly folded into cristae (increases SA) -contains transport proteins and redox-performing proteins (forming electron transport chain) -contains enzymes involved in energy production

Answer 278

-have porins (large channels) which allow molecules <5000kDa to enter -contain enzymes for mitochondrial lipid synthesis

Answer 279

29S and 28S ribosomes

Answer 280

double stranded circular chromosome about 15-17kbps long -encodes 37 genes -inherited from mother

Answer 281

-can't be newly synthesised so arise from existing mitochondria via fission

Answer 282

-debris is segregated -fission (mitochondria splits into two) -fusion (occurs as a stress response -two mitochondria fuse to form one) -mitophagy (mitochondria is engulfed into an autophagosome and is degraded) -biogenesis

Answer 283

-TOM (translocator of outer membrane) -TIM (translocator of inner membrane) -SAM (sorting and assembly machinery) -OXA (cytochrome oxidase activity)

Answer 284

-proteins are already synthesised (interacting proteins like chaperones are bound to synthesised proteins to stop them folding before they are docked to TOM) and then translocated into the mitochondria using signal sequencing and translocation proteins (eg. TOM, TIM, OXA, SAM) --protein precursor binds to receptor protein in TOM complex -it is then inserted into the membrane by the TOM complex and translocated into the inter-membrane space and then translocated through the TIM23 complex into the matrix (goes through TOM and TIM at same time) -polypeptide signal is cleaved off by signal peptidase

Answer 285

-chaperones bind to newly synthesised proteins, to stop them folding before they get translocated by TOM -chaperones must then be dissociated from the polypeptide chain, using ATP

Answer 286

-most common: using TOM and TIM or -protein completely enters matrix -signal sequence is cleaved, which unmasks a second sequence, causing insertion into OXA (cytochrome oxidase activity) complex

Answer 287

-issue: TOM can’t insert proteins into bilayers -instead, chaperones keep proteins (typically porins, which are beta-barrel proteins) unfolded as they enter the intermembrane space -SAM complex inserts the proteins into the outer mitochondrial membrane and folds them

Answer 288

vesicles found in all eukaryotic cells which carry out oxidation and reduction rxns

Answer 289

-surrounded by a single membrane -have no DNA or ribosomes

Answer 290

carry out oxidative rxns (remove hydrogen atoms from inorganic compounds) eg. b-oxidation of fatty acids, detoxification of alcohol

Answer 291

produced by endoplasmic reticulum -peroxisomal precursor vesicle buds off from ER -peroxisomal precursor vesicle uptakes peroxisomal proteins and lipids from cytosol and grows, becoming a peroxisome or from pre-existing peroxisomes -peroxisome grows -fission occurs, producing two daughter peroxisomes

Answer 292

-Pex5 recognises signal sequence (Ser-Lys-Leu) and accompanies cargo into peroxisome, is ubiquitylated and cycled back into cytosol

Answer 293

severe brain, liver and kidney abnormalities caused by a mutation in Pex5 gene (gene involved in translocating proteins into peroxisomes)

Answer 294

-energy -directions -mechanical interactions (w/something outside cell) -microtubules/actin microfilaments

Answer 295

hollow tubes of alpha and beta tubulin

Answer 296

-13 protofilaments -24nm diameter -tubulin dimers of alpha and beta tubulin

Answer 297

-cells, protozoa, sperm, etc -moving fluids, reproductive tracts, etc -using cilia/flagella

Answer 298

different lengths (cilia shorter than flagella)

Answer 299

same structure -both have an axomere

Answer 300

9+2 microtubule arrangement -9 outer doublets consisting of complete fibres (13 protofilaments), incomplete fibres (10 protofilaments) and dynein arms and radical spokes -inner pair

Answer 301

dynein arms on one side have contact but don't have contact on other side

Answer 302

nexin crosslinks between doublets

Answer 303

via power strokes and recovery strokes -controlled by outer dynein arms

Answer 304

wave-like waveform -controlled by inner dynein arms

Answer 305

control waveforms of cilia and flagella -inner arm controls waveform -outer arm controls power

Answer 306

continuation of axoneme inside cell -shorter than axoneme

Answer 307

9x3 microtubule array

Answer 308

-actin is disassembled at - end -actin removed is recycled (by ATP being added) by being added to the + end (causing movement in the direction of the + end)

Answer 309

-microtubules are wider -actin filaments are solid whilst microtubules are hollow -actin filaments have homomeric (identical) subunits but microtubules have different subunits (alpha and beta) -actin filament subunits are 42kDa but microtubule subunits are 50kDa -actin filaments have 375 amino acids whist microtubules have 450 -actin filaments have ATP in monomer (hydrolysed to ADP during assembly so ADP in filament) but microtubules have GDP in monomer (GDP in alpha, DTP in beta)

Answer 310

-monomers added to + end -ATP hydrolysed to ADP

Answer 311

control movement (inhibit by binding ATP eg. profilin or activate)

Answer 312

-sequence monomers of actin filaments -nucleate monomers -polymerise monomers to produce actin filaments -cap actin filaments (end-blocking) -cross-link actin filaments -bundle actin filaments -severe actin filaments -bind actin filaments to membrane -depolymerise actin filaments (back into monomers)

Answer 313

microtubule

Answer 314

-Ca2+ binds to troponin, causing it to move and reveal the active site -cross-link forms between myosin and actin -myosin head bends, releasing ADP and Pi -new ATP binds to myosin and cross-link breaks -ATP is hydrolysed to ADP and Pi, returning myosin to its original position

Answer 315

-signal stimulates filopodium -lamellipodium pushes cell forwards -new adhesions form with the matrix -old adhesions at back of cell with the matrix break -cell contracts and moves forward -cycle continues

Answer 316

extracellular meshwork of proteins and hydrated macromolecules (secreted)

Answer 317

-bones (calcified) -keratin (fibrous and loose) -tendons (tensile strength) -skin (elastic) -cornea (transparent)

Answer 318

-regulate migration -regulate tissue integrity and cell shape -regulate proliferation (checks cells and encourages independent growth)

Answer 319

-fibrous proteins (collagen, elastin) -adhesion proteins (fibronectin, laminin) -hydrated sugars

Answer 320

-repetitive structure: Gly-Pro-Hyp triplet repeat -triple helix -3 alpha chains

Answer 321

glycine-proline-hydroxyproline

Answer 322

fibroblasts and epithelial cells

Answer 323

defect in collagen

Answer 324

-in a multi-step process driven by self-assembly -uses vitamin C as cofactor -involved cleavage of pro-peptides and assembly of fibrils

Answer 325

lysyl oxidase

Answer 326

stretches, revealing single elastin molecules and cross links

Answer 327

defect in fibrillin

Answer 328

-repetitive disaccharide chains 70-200 units long -highly sulphated (charged) -often substituted into proteins using a link tetrasaccharide

Answer 329

proteoglycans and glycoproteins

Answer 330

-laminin -fibronectin -integrins -focal adhesions

Answer 331

-cruciform structure -heterotrimer (β, α and γ chains) -have globular domains and coiled-coil-α-helical domain

Answer 332

-disulphide crosslinks -collagen binding site -cell binding site (type 3 receptors)

Answer 333

-heterodimer (α and β subunits non-covalently associated) -α subunit with divalent cations bound -β subunit with cysteine rich domains -matrix binding site -COOH groups in cytosol

Answer 334

bind matrix through divalent cations -removing divalent cations causing cell to detach)

Answer 335

-integrin -cytoskeletal proteins eg. talin

Answer 336

form mechanical links between intracellular actin and extracellular myosin -involved in signalling -transmembrane receptor

Answer 337

-platelets binding fibroniogen to clot blood -Glonemann's thrombasthenia -bleeding gums/noses -leukocyte adhesion deficiency (LAD) syndrome -impaired expression -recurrent bacterial infections

Answer 338

-cell-matrix anchoring junctions -cell-cell anchoring junctions -tight junctions -channels forming junctions

Answer 339

transmembrane proteins which mediate cell-cell adhesion -Ca2+ dependent

Answer 340

-bundle of actin filaments -contains cadherins -links to cytoskeletan

Answer 341

-hemidesmosomes attach cells to basal lamella but desmosomes attach cells to cells -hemidesmosomes contain integrins but desmosomes contain cadherins (integrins and cadherins link to intermediate filaments)

Answer 342

between cells

Answer 343

between cell and basal lamina

Answer 344

an autoimmune disease where skin blisters due to failure in desmoglein (α cadherin) which holds keratinocytes in epidermis

Answer 345

between cells in clusters near apical face

Answer 346

electron dense dye

Answer 347

transport of substances across epithelium by passing through intercellular space (gap junctions) -passive -regulated

Answer 348

transport of substances across epithelium by passing through transmembrane proteins in the apical and basal membranes and the cytoplasm in between -active

Answer 349

between cells -patches rather than continuous barrier -found in connective tissue, neurones, heart muscle, epithelia (places where regulation is key)

Answer 350

-6 subunits called connexins (can be homomeric or heteromic) -flexibility in arangements allows gap junctions to be regulated

Answer 351

-pH -membrane potential -Ca2+ -cell signalling

Answer 352

-chromosomal replication -chromosomal segregation -cell division

Answer 353

when stimulated by extrinsic factors eg. nutrient status, integrins, TGF-β receptors, G-protein coupled receptors, tyrosine kinase receptors -these can be overruled by signalling proteins (which force proliferation to stop)

Answer 354

nutrient status -integrins -TGF-β receptors -G-protein coupled receptors -tyrosine kinase receptors

Answer 355

by signalling proteins

Answer 356

the way a cell decides whether to proliferate or not based on signals from environment -decides cell fate

Answer 357

-cells increase in size -ribosomes and RNA are produced -preparation for DNA synthesis

Answer 358

-DNA synthesis (chromosomes duplicated)

Answer 359

-fidelity of DNA is checked -preparation for nuclear division

Answer 360

-mitosis (prophase, prometaphase, metaphase, anaphase, telophase) -cytokinesis

Answer 361

-interphase (G1, synthesis and G2) -mitosis and cytokinesis

Answer 362

-end of G1 -during synthesis -during G2 -during mitosis

Answer 363

DNA damage

Answer 364

DNA damage and stalled replication forks

Answer 365

damaged/unreplicated DNA

Answer 366

correct attachment of chromosomes to spindle and correct separation of chromosomes to poles of cell

Answer 367

-to stop cell cycle if there's a problem -to allow for increase of scheduled lengths of phases -to facilitate repair processes

Answer 368

-if serum and growth factors are removed before cells have completed 80-90% of G1, they revert to G0 (don't continue with cell cycle) -if serum and growth factors are removed during the final hour of G1, they proceed to synthesis, etc

Answer 369

-cyclin -cyclin dependent kinases (cdk) -diff cyclins and diff cdks needed for diff phases

Answer 370

-genetic approach: looking for mutations (studying mutant cells that can't proceed into next phase of cell cycle) -biochemical approach: looking for proteins involved in transition (studying large no. cells undergoing same transition at same time)

Answer 371

-is obvious by looking at yeast cells which phase of cell cycle they are in -have fast division rate -cell cycle genes are highly conserved -yeast cells can be haploid or diploid

Answer 372

-deplete cytoplasm of diff proteins -remove cytoplasm at diff stages to study changes over time -protein electrophoresis can be carried out

Answer 373

-kinases phosphorylate target proteins to turn them on (using ATP) -phosphatases dephosphorylate phosphorylated target proteins to turn them off

Answer 374

-extract is incubated with ATP and substrate -electrophoresis of substrate is carried out

Answer 375

linear DNA molecule

Answer 376

chromosomes with the same genes in the same order -one from mother, one from father

Answer 377

newly copied DNA strands still joined together by centromere

Answer 378

sister chromatids condense

Answer 379

mitotic spindle attaches to kinetochore by microtubules

Answer 380

sister chromatids separate to opposite poles of the cell

Answer 381

nuclear envelope reassembles

Answer 382

M-cdk complex (using cyclinB)

Answer 383

-assembly of mitotic spindle -attachment of each sister chromatid to opposite poles -condensation of chromosomes -breakdown of nuclear envelope -rearrangement of cytoskeleton and golgi body

Answer 384

-M-cyclin binds to cdk 1, forming inactive M-cdk -cdk-activating kinase (CAK) and active phosphatase (cdc25) activate M-cdk by removing inhibitory phosphate

Answer 385

-promote the activation of phosphatase (cdc25) -inhibit the cdk-inhibitory kinase (Wee1) -this forces the switch from G2 to M phase -increase in cyclinB expression increases the levels of M-cyclin

Answer 386

M-cdk complex

Answer 387

Anaphase-promoting complex (APC) -a ubiquitin ligase (promotes ubiquitination and degradation) -causes the degradation of M-cyclin in proteasome and securin

Answer 388

-S and M cyclins (if they are degraded, most cdk targets will be dephosphorylated by phosphatase, which inactivates the cdks) -securin (if degraded, securin can no longer protect protein linkages holding sister chromatids together and a protease is activated that separates the sister chromatids)

Answer 389

chromosomes end up in wrong daughter cell (chromosome non-disfunction) -causing mutations on both alleles, resulting in a phenotypic change (2 hit hypothesis)

Answer 390

-astral microtubules -interpolar microtubules -kinetochore microtubules

Answer 391

centriole surrounded by pericentriolar matrix (highly structured matrix containing lots of proteins) to nucleate the microtubules

Answer 392

microtubules released from centrosome which have contact with cell cortex to position the spindle

Answer 393

microtubules which attach to the chromosomes at kinetochore

Answer 394

microtubules which overlap with the interpolar microtubules from the opposite spindle pole

Answer 395

when attached correctly, kinetochore is pulled in opposite directions but sister chromatids resist -creating tension (which is sensed)

Answer 396

when attached incorrectly, tension is lower so inhibitory signal is released, causing microtubuial attachment to loosen

Answer 397

correctly -tension (from kinetochores being pulled in opposite directions) incorrectly -less tension

Answer 398

securin is degraded

Answer 399

cleaves and dissociates the cohesins between sister chromatids, allowing them to separate as the kinetochore microtubules shorten (allows anaphase to occur)

Answer 400

kinetochore microtubules shorten, pulling chromatids apart and breaking the cohesin bridges

Answer 401

-astral microtubules pull poles of cells further apart by motors and depolymerisation -interpolar microtubules slide past eachother

Answer 402

-loss of heterozygosity by non-disfunction (chromatids separated unevenly) -loss of heterozygosity by recombination (chromosome arms swap) -loss of heterozygosity by gene conversion (polymerase jumps across close strands and replicates some of other template strand)

Answer 403

-chromatids separated unevenly so that one daughter cell has more (3/1) -this can be eliminates by apoptosis -however, if it isn't, cell may try to divide again and extra chromosome may be lost -resulting in hemizygous cell (with only one chromosome) where no active protein can be made

Answer 404

-arms of chromosomes swap, causing mutant alleles to be on diff arms -separation works but could result i a cell with 2 mutated alleles

Answer 405

if strands are too close together, polymerase could jump across the strands and replicate some of other template strand

Answer 406

-maternal and paternal homologues pair up -genetic diversity is generated by regeneration between homologous chromosomes -1 complete chromosome is pulled apart

Answer 407

-separation into haploid daughter cells

Answer 408

-homologues pair up -pairing facilitated by synaptonemal complex

Answer 409

X-shaped site of chromosomal recombination in paired homologues during meiosis

Answer 410

Complex formed when homologues pair up -paired homologues bought 400nm apart -complex formed by axial cores being cross-linked by transverse filaments -complex aids binding -zips replicated DNA together

Answer 411

axial cores (proteins that bind chromatin using cohesion) are cross-linked by transverse filaments

Answer 412

The observation that when one crossover is forming, other crossovers nearby are inhibited from forming -limits how much crossing over can occur

Answer 413

-chromosome number -structural arrangement of chromosomes

Answer 414

disorder of chromosome number -caused by chromosome non-disfunction -monosomy, trisomy, polyploidy, etc

Answer 415

by Spectral Karyotyping (SKY)

Answer 416

-when it occurs in meiosis 1, all the gametes are affected -when it occurs in meiosis 2, only half of the gametes are affected

Answer 417

a type of aneuploidy where one chromosome is missing (only have one copy of chromosome) -embryonically lethal

Answer 418

a type of aneuploidy where there are three copies of chromosomes

Answer 419

a type of aneuploidy where there is a whole extra set of chromosomes

Answer 420

-embryonically lethal (eg. triploid -XXX/XYY/XXY, nullsomy -missing chromosome pair, monosomy) -minor issues (eg. additional sex chromosome) -infertility (eg. missing sex chromosome -X)

Answer 421

during homologous recombination

Answer 422

via necrosis (uncontrolled) or apoptosis (controlled)

Answer 423

-physical damage (extreme temps, cuts, burns) -toxins (internal or external) -acute hypoxia (low oxygen concs) -ischaemia (loss of blood supply)

Answer 424

-physiological situations (tissue cell maintenance, developmental cells loss, immune response, hormone-dependent involution) -pathological situations (DNA damage, virally infected cells)

Answer 425

-compromised membrane integrity -swelling of organelles/cells -increased intracellular calcium -actolysis (enzymes start destroying their own cells) -cell lysis (bursting)

Answer 426

-shrinkage -nuclear breakdown -apoptic bodies (vesicles containing dying parts of cells) -phagocytosis triggered -energy required (apoptosis is an active process)

Answer 427

no -because it is controlled

Answer 428

yes -triggered by cell lysis (bursting)

Answer 429

-when cells in middle start dying via necrosis -detected by body -so cells surrounding them die via apoptosis to limit the spread of necrotic cell death eg. in brain ischaemia

Answer 430

shrinks/condenses

Answer 431

maintained

Answer 432

packaging into apoptic bodies

Answer 433

leakage to intracellular fluid

Answer 434

fragments chromatin condenses

Answer 435

genes involved in apoptosis -recognise apoptotic signal -phagocytosis of apoptotic cells

Answer 436

BH3 (EGL-1) Bcl-2(Ced 9) APAF-1 (Ced 4) Caspases (Ced 3)

Answer 437

ced genes which carry out cell death (essential for apoptosis) -initiators or executioners -irreversible pathways

Answer 438

-has a cysteine residue in active site -are aspartic acids -cleavage site in target protein

Answer 439

caspases activated by apoptotic signals which activate executioner caspases

Answer 440

caspases which cleave over 1000 proteins

Answer 441

one initiator caspade can activate multiple executioner caspases

Answer 442

-cause breakdown of nuclear structure (eg. cleave nuclear lamins to breakdown lamella) -prevent DNA repair by cleaving poly ADP-ribose polymerase (PARP) -cause cytoskeletal changes (eg. cleave cytoskeletal proteins to breakdown actin)

Answer 443

-extrinsic pathway -intrinsic pathhway

Answer 444

-apoptic signal (eg. ligand on lymphocyte) binds to Fas death receptors on cell -this causes the death-induced signalling complex (DISC), which composes of caspases and FADD adaptor proteins, to assemble -caspases are activated and cleaved -executioner caspases cause the cell to undergo apoptosis

Answer 445

-apoptic signal causes mitochondria to release cytochrome c -cytochrome c activated Aparf1 (apoptic protease activating factor) -apoptosome assembles -caspase 9 is recruted -caspase 9 is activated and cleaved -executioner caspases are activated and cause the cell to undergo apoptosis

Answer 446

-stress signals (eg. DNA damage) -developmental damage

Answer 447

death ligands

Answer 448

disease of aberrant cell proliferation and differentiation

Answer 449

by studying people migrating -cancer trends fitted with place they moved to

Answer 450

-infection -diet -noxious substances

Answer 451

using cytogenetic analysis -condensed chromosomes from cells undergoing mitosis -identify abnormalities s -use molecular probes and FISH

Answer 452

abnormality in chromosome 22 of leukaemia cells in chronic myeloid leukaemia -chromosome translocation

Answer 453

-ABL (a protein kinase) can not switch itself off -causes constant proliferation

Answer 454

-oncogenes -tumour suppressor genes -DNA-repair genes

Answer 455

genes with the potential to cause cancer by transforming cellular behaviour to cause proliferation -generally dominant

Answer 456

-deletion/point mutation in coding sequence -hyperactive protein produced -regulatory mutation -more of normal protein produced -gene amplification -normal protein is overproduced -chromosome rearrangement -hyperactive fusion protein produced or normal protein is overproduced

Answer 457

-first human oncogene identified -small GTPase (on -RasGTP, off -RasGDP) -important in growth factor-induced growth

Answer 458

Ras is only in active form (GTP-bound form) -looses ability to hydrolyse GTP to GDP -causes cell to growth without any control

Answer 459

-tyrosine kinases dymerize in presence of growth factor -take up Grb2 and Sos -this turns Ras on -Ras turns diff pathways on eg. cell growth, gene expression, cell movement

Answer 460

-normal cell and cancerous cell are fused -hybrid cells are inserted into immune-compromised mouse -no tumours formed in mouse ∴normal genes must be dominant to cancerous (normal cells must expressor tumour suppressor genes, which get lost in oncogenesis)

Answer 461

gene which helps to prevent cancer formation -recessive genes -mutations in tumour suppressor favour cancer formation

Answer 462

more than one (one is not sufficient) -5-8 driver mutations

Answer 463

have an increased tendency for mutations

Answer 464

-can visualise chromosomal translocation -see changes in copy number, interchromosomal rearrangement, intrachromosomal rearrangement

Answer 465

-DNA repair pathways -correction mechanisms for DNA repair errors -mechanisms for DNA segregation errors

Answer 466

tumorigenesis

Answer 467

tumorigenesis

Answer 468

though p53 signalling pathway -stable, active p53 causes cell cycle arrest, senescence and apoptosis

Answer 469

cell cycle checkpoint gene involving in the signalling pathways in response to cellular stresses (eg. DNA damage, telomere shortening, hyperproliferative signals)

Answer 470

is mutated -disrupts instrinsic apoptosis

Answer 471

-disrupts intrinsic apoptosis -contributes to genetic instability

Answer 472

enzymes with a RNA catalytic subunit -involved in RNA splicing

Answer 473

-receives incoming vesicles -sorts internalised cargo (either for degradation or to return to membrane) -develop into late endosomes (for degradation)

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MBB11003 -Molecular and Cell Biology Flashcards

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