Brainscape's Knowledge GenomeTM


Top Flashcards (Certified)
All Flashcards

MCB - Molecular Bio > prokaryotic molecular biology > Flashcards

prokaryotic molecular biology Flashcards

(107 cards)

Start Studying
1
Q

prokaryotes features and genome organisation

A

bacteria + archaea
absence of membrane-bound organelles

DNA in nucleoid

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
2
Q

most DNA in B form (like regular double helix)

A

2 polynucleotide chains in opposite orientations (run in opposite directions)
regular RH double helix
2nm diameter
complete turn ever 3.4nm
10.5 base pairs per turn helix
flexible - in terms of base pairs per turn, 3D gelix, bends

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
3
Q

supercoiling

A

in circular DNA
double helix has additional turns or turns removed
adds tension and torsional stress

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
4
Q

negative supercoiling

A

twist opposite to RH turn of helix

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
5
Q

positive supercoiling

A

twist same direction as turn of RH helix

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
6
Q

torsional stress

A

by formation superhelices or alter no. base pairs per turn (change amount of twisting)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
7
Q

linking number

A

how tight the twist (L)

total no. times 2 strands of helix of closed molecule cross each other when constrained to lie in a plane

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
8
Q

topoisomerases

A

enzymes that alter linking number of DNA molecule

type I and II

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
9
Q

type 1 topoisomerase

A

break 1 strand, pass other strand through gap and seal break
L changed by +/- 1
reduce coils so looser and easier for transcription/translation

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
10
Q

type 2 topoisomerase

A

breaks both strands, pass another part of helix through
L changed by +/- 2
removes stress and can replicate both strands at the same time

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
11
Q

E.coli genome organisation

A

supercoiled loops radiate from central protein core

proteins in nucleoid: DNA gyrase, topoisomerase 1, 4 heat unstable (HU) proteins for packaging DNA
forms tetramer around DNA (like histones in eukaryotes)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
12
Q

nucleoid

A

(meaning nucleus-like) is an irregularly-shaped region within the cell of a prokaryote that contains genetic material, not surrounded by a nuclear membrane like eukaryotes

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
13
Q

different prokaryote DNA

A

not all have circular DNA
multipartite - divided into 2 or more DNA molecules (or might be plasmid)
sizes vary (obligates have smaller genomes)
min no. genes is 200 but obligate have 120

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
14
Q

horizontal gene transfer

A 
rapid changes important in evolution
prophages - genomes with phage-like elements linked to pathogenesis
genomic islands (GIs) often mutated so pathogenesis
transposable genetic elements can change position in genome
How well did you know this?
1
Not at all
2
3
4
5
Perfectly
15
Q

main replicative enzyme

other enzymes in replication

A

polymerase III

pol I removes RNA primers from Okazaki
ligase links fragments

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
16
Q

replicon

A

molecule that replicates from a single origin of replication
basic unit of replication
single one for each genome
DNA molecule/sequence replicated at least once per cell division cycle

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
17
Q

E.coli replication features

A

oriC single origin of replication (replicon)
bidirectional replication leads to theta (egg with line through) structure
DNA digested with restriction enzymes and ligated to plasmid lacking origin of replication

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
18
Q

E.coli replication process

A

1) initiation - 20 monomers DnaA bind to 4/5 copies of 9bp sequence in RH 2/3 (2 thirds) of oriC, DNA bends and forms closed complex
2) 3 copies of AT-rich 13bp sequence on LH 1/3 (a third) of oriC –> melt so open complex and separate DNA strands
3) DnaB helicase loaded onto melted DNA w/ DnaC, ATP hydrolysed so DnaC released
4) DnaB unwinds DNA in both directions which requires SSB protein (protects ssDNA) and DNA gyrase
5) RNA pol III makes DNA, RNA primer on both strands (primase)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
19
Q

control of E.coli replication

A

Dam methylase - methylates adenine residues GATC in OriC - replication only initiated if all 14 methylated

hemi-methylated replication because new strand unmethylated and can’t replicate so them re-methylated 1/3 through cell cycle

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
20
Q

E.coli replication termination

A

2 forks approach each other and fuse in terminus region opposite oriC
arrest forks from 1 direction (polar)
Tus protein stops fork and waits for other form, if no other fork then carries on to next fork (ensures terminate both same time)

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
21
Q

why is E.coli termination regions further apart than B.subtilis

A

because makes DNA as fast as can so spread them out in case 1 fork slower

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
22
Q

recombination

A

breaking and rejoining DNA to new combinations for diversity and DNA repair

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
23
Q

homologous recombination (definition + process)

A

2 sequences have similarity

1) alignment - of homologous regions
2) cleavage - of 1 strand at chi sites (RecBCD endonuclease)
3) invasion - 1 cleaved strand invades other sequence of 2 strands, Holliday junction (RecA), joined
4) branch migration - slide along and increase heteroduplex (RuvAB) so spread branch
5) isomerization - DNA twist, junction cross and uncross, chi form
6) resolution - crossed strands cleaved (RuvC), end depends on horizontal or vertical cleavage

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
24
Q

non-homologous recombination

A

lack sequence similarity

How well did you know this?
1
Not at all
2
3
4
5
Perfectly
25
RecBCD
binds end of dsDNA and unwinds with helicase activity degrades both ssDNA strands with dual5' to 3' and 3' to 5' exonuclease activity encounters chi site - 3' to 5' exonuclease inhibited, 5' to 3' stimulated so degrades 5' end not 3' ssDNA 3' end tail and RecA binds to this
26
transposition (3 main points)
genes change position on a chromosome transposable genetic elements Tn3 family Inversion sequences
27
transposable genetic elements | definition, classes
sequences that can transfer copies of themselves to other parts/other DNA/other orientation 7 classes in bacteria: insertion+composite, Tn3, phages, inversion, Tn7, Gram +ve
28
insertion sequences
``` small (<2kb) encode proteins for transposition bound by inverted repeats cause direct repeats at site of insertion insert anywhere or specific target sites ```
29
transposons
Tn3 family bigger encode phenotypic characteristic like drug resistance
30
Tn3 family structure
lecture 2 page 1 diagram (IR, tnpA, res, tnpR, Bla, IR) 38bp terminal repeats (IR) encodes 3 proteins TnpA + TnpR + Bla (beta-lactamase=ampicillin resistance) 3 cis-acting elements internal resolution site (IRS/res)
31
Tn3 family replicative transposition
formation of cointegrate - 2 transposon copies in direct repeat occur at junction of component molecules (transposase) recombination between 2 res sites forms 2 molecules each with res site (resolvase) lecture 2 page 1 diagram
32
Tn3 family non-replicative transposition
conservative Tn10 + other transposons (leaves no copy behind, just 1 molecule)
33
inversion sequences
alter orientation within DNA | control gene expression
34
salmonella spp inversion
2 types of flagellum change to avoid immune system phase 1/2 H antigen determined by orientation of hin region (H-inversion) hin region - bound by 2 IR, encodes invertase H1 has own promoter and operator and separated from hin region H2 is an operon with rep gene which suppresses H1 and promoter within hin hin faces certain way for promoter to point towards H2 so rep repressed H1 (phase 2) if towards H1 then p for H2 in wrong orientation so H1 expressed (phase 1)
35
DNA damage
change from normal nucleotide sequence and supercoiled double helical state from physical/chemical env. and errors in replication single base changes structural distortions
36
single base changes
no effect on transcription/replication e.g. keto-enol tautomerisation, deamination of cytosine to uracil, U rather than T, chem mods
37
structural distortions
may impede transcription/replication e.g. single strand breaks, covalent modification of bases, removal of base, inter/intra strand covalent bonds, thymine dimer formation
38
thymine dimer formation
intrastrand binding from UV so won't divide properly | 2 adjacent thymines on same strand covalently link in cyclobutaine structure/ 6-4 photoproduct
39
DNA repair (5)
``` direct mismatch excision tolerance retrieval ```
40
direct DNA repair (and e.g.)
reversal/removal photolyase: photoreactivation repairs UV-induced dimer to create 2 thymine residues again deoxyribopyrimidine photolyase enzyme contains 2 chromophores which absorb light and splits cyclobutane structures
41
mismatch DNA repair (and e.g.)
detect and repair mismatched bases Uracil DNA glycosylase: when U instead of T, removes U making AP site then AP endonuclease breaks phosphodiester backbone at site and DNA Pol 1 binds and lays new DNA, gap sealed by DNA ligase mut system: Mut S recognise mismatch/insertion/deletions (indels) and binds MutL stabilise complex Mut S-Mut L activate Mut H which locates methyl group and nicks opposite because new strand more likely wrong Mut U helicase II unwinds DNA from nick to mismatch DNA Pol 1 degrades and replaces DNA, ligase seals
42
excision DNA repair (and e.g.)
large bit of DNA replaced by undamaged (knows damage is somewhere) E.coli 3 modes: very short patch, short 20, long 1500-10000bps uvrABC enzyme bind damage and incision on both sides UvrD separates strands DNA Pol 1 and ligase
43
tolerance DNA repair (and e.g.)
replication proceed through damage replace and guess what goes next low-fidelity (incomplete) DNA pols synthesise past damage (almost all Y-family) E.coli: pol IV and V humans: 5 pols, pol n (eta) bypass UV photoproduct, defective in xeroderma pigmentosum (variant of skin genetic disorder) so helps prevent UV cancer
44
retrieval DNA repair (and e.g.)
recombination with other copies of DNA daughter strand gap repair: takes good bit from other DNA copy to replace wrong, then rely on other repair like excision, not actually repair itself and only when severe damage
45
AP site
apyrinic/apyrimidinic
46
SOS response
E.coli activates DNA repair genes if severe damage LexA protein represses expression of SOS operons (LexA binds to LexA box in promoter of genes) RecA changes conformation to active to induce SOS, inactivates LexA inhibits 3'-5' editing in DNA Pol III allowing error-prone replication sulA inhibits cell division
47
transcription overview
template recognition - RNA pol bind at promoter initiation - 9 internucleotide phosphodiester bonds, sigma factor release elongation - addition of nucleotides termination - dissociation of enzyme, RNA and DNA rho-dependent/independent
48
RNA Pol
complete holoenzyme is a2bb'ωσ σ factor easily dissociate to form core enzyme
49
holoenzyme
enzyme with coenzyme
50
factors controlling gene expression
``` promotor recognition promoter strength alternative σ factors guanosine tetraphosphate (iinhibits RNA synth.) mRNA degradation RNA processing regulatory RNAs regulate transcription initiation ```
51
promoter recognition
core enzyme bind many sites in loose closed complex with 1hr dissociation half life holoenzyme loosely bound in closed complex has dissociation half life of 1 second holoenzyme tightly bound in open complex has dissociation half life of >1hr so σ makes RNA pol specific for promoters, tight stays for long time and melts DNA to open it
52
conserved promoter sequence
TATAAT - 10 Pribnow box | TTGACA - 35
53
UP mutations
make promoter more similar to consensus
54
consensus sequence
a sequence of DNA having similar structure and function in different organisms.
55
DOWN mutation
make less similar to consensus mutations in -35 region reduces rate of closed complex formation (initial promoter recognition) in -10 region reduces rate of open complex formation (required for melting)
56
UP elements and their recognition
AT rich region upstream of -35 region which affects promoter strength (affects rate of transcription initiation) alpha subunits involved in UP element recognition: 2 independently folded domains (alpha-NTD and alpha-CTD) cleaved a-CTD can still dimerize and bind DNA and interacts with DNA of UP element RNA Pol without a-CTD assemble and transcribe but no enchanced activity from UP
57
initiation
release of σ factor increases affinity of RNA Pol for non-promoter DNA, change of shape locks Pol to DNA
58
alternative σ factors
transcribe specific subsets of genes (heat shock, motility, nitrogen metabolism)
59
constitutive expression
some genes that are always expressed | most genes are not constitutive but they are regulated and controlled
60
operon
some genes transcribed together with single promoter and is regulated as a single gene with a linked function polycistronic mRNA (more than 1 gene per mRNA)
61
what are 2 transcription mechanisms that control gene expression?
induction - switching genes on when required, by activators (apoinducers) repression - switching genes off by reressors
62
regulatory proteins like repressors are activated by
small molecules called effectors
63
inducers
activate the activators like apoinducers so turn genes on
64
co-repressors
activate repressors or inactivate activators so turn genes off
65
regulon
group of associated (by physiological function) genes, that may not be in the same operon but are controlled by a single regulatory protein e.g. PHO in E.coli
66
global regulation
a single environmental factor causes regulation of many genes with diff metabolic functions like SOS response which has 30 genes
67
lac operon structure
polycistronic RNA with 3 protein coding genes lacZ lacY lacA and lac I, lac P, lac O, CAP site
68
polycistronic
encode more than one polypeptide separately within the same RNA molecule
69
describe the lac operon sugar metabolism process
E.coli uses diauxic growth which is using 2 substrates in succession not together if glucose is available it is used first and it represses lactose enzymes (catabolite repression) in the lag phase when glucose runs out, it induces lactose enzymes b-galactosidase enzyme makes allolactose (lactose isomer) which binds to each 4 subunits of repressor this causes an allosteric shape change so it can't find the operator IPTG binds repressor and activates promoter no glucose activates adenyl cyclase to convert ATP to cAMP which activates CAP protein which binds CAP site and recruits RNA Pol to promoter
70
lacZ
beta-galactosidase cleaves b-galactosides into monosaccharides
71
lacY
beta-galactoside permease transports b-galactosides into cell
72
lacA
beta-galactoside transacetylase detoxify b-galactosides by acetylation
73
lacI
repressor gene, always expressed | 182bp up from lacZ and p/o/CAP
74
lacP
promoter | where RNA Pol binds
75
lacO
operator overlaps promoter where lac repressor binds so blocks promoter
76
CAP site
where cAMP acceptor protein binds
77
how does CAP interact with RNA Pol
alpha subunit of Pol interacts with 7AA surface-exposed beta-turn of CAP
78
trans-acting elements
diffusable products regulate gene expression on genes distant from where transcribed
79
cis-acting elements
regulate genes on DNA encoded on
80
complementation (and E.coli example)
if 1 on 1 chromosome working then normal phenotype
81
lac mutations (experiment with merodiploid E.coli so 2 copies or genes)
functional proteins (Z,Y,A) are still transcribed because mutated genes complemented by other chromosome lacI still transcribed when 1 chromosome has mutated gene but there are some dominant mutations as well that overpower the other chromosome so mean the non-functional protein is transcribed that interferes with ability to bind operator lacO and lacP binding site protein mutations are not transcribed
82
cloning
insertion of a DNA fragment into a self-replicating element to copy/isolate a particular piece of DNA
83
process of cloning
1) vector and DNA are restricted to make sticky ends which bind together and the gap is sealed with DNA ligase to make a circular recombinant plasmid 2) make competent with stresses like chemicals so can import into cell 3) select for plasmid-containing cells with selectable marker like antibiotic resistance so only cells with plasmid survive and a vector screenable marker to determine plasmid contains DNA and not just vector with plasmid
84
why is cloning used?
to analyse mRNA transcripts | it's made into cDNA and then its RNA is cloned so reverse transcription
85
vector types
``` phage insertion vectors replacement vectors fosmids/cosmids Ti plasmid BACs YACs ```
86
phage vector
produce large numbers and incorporate DNA into chromosomes | can replace non-essential genes with cloned DNA
87
insertion vectors
non essential DNA already removed at restriction site | DNA cloned in
88
replacement vectors
non essential DNA replaced with non-coding stuffer DNA which can then be replaced by DNA to be cloned
89
fosmids/cosmids
large plasmids that clone large fragments | based on F-plasmids with co-sites (to help package vector into phage) and lacZ and T7 promoter
90
Ti plasmid
found in pathogen, used to transfect plant cells (but take pathogenic bit out beforehand)
91
BACs
bacterial artificial chromosomes very large fragments based on F-plasmid, carry selectable markers and lacZ gene
92
YACs
yeast artificial chromosomes based on real yeast chromosomes, grow in yeast cells must contain origins of replication, centromere, telomeres telomeres are joined and circularised to be stable carry 2 selectable and 1 screenable marker transfected into mutant yeast which is trp1- on one arm and ura3+ on the other arm and must contain both arms for growth the cloning site is within the sup4 gene
93
what do eukaryotes use to regulate gene expression?
small non-coding RNA (ncRNA) | e.g. addiction cassette in plasmids
94
1 type of addiction cassette
makes stable mRNA of lethal membrane protein and also makes an anti-sense ncRNA which is unstable and binds to the mRNA and prevents translation plasmid with ncRNA can pass to daughter cell in replication and so repress lethal mRNA (if no plasmid then dies from lethal mRNA)
95
hok sok system
host killing system by the R1 plasmid in E.coli is an example of an addiction cassette
96
RyhB
ncRNA involved in controlling iron use in E.coli E.coli has proteins that need iron but they are not essential so when iron is limited FUR (global regulator) stops repressing ryhB which binds mRNA or non essential iron proteins and degrades them so the iron need falls
97
positive interactions in microbial communities
toxic product of 1 organism may be substrate for another | e.g. cyanobacteria needs heterotrophic bacteria
98
Windogradsky columns
mixed soil/sediment/waater/carbon/nutrients and leave for years colours form from bacterial growth, whole ecosystem forms
99
negative interactions
Ab production targets others, competition for substrates
100
Quorum sensing
similar organisms send chemical signals and molecules like autoinducers (in response to cell population density changes) and sense each other once threshold of autoinducers is reached, expression and function is changed can use to create biofilm
101
S. aureus quorum sensing process and effects
forms biofilms in wounds and medical devices and quorum sensing drives the switch from biofilm to invasive with the Agr regulatory system at low density AIP is low so AgrC is inactive so no RNA III and no response so biofilm forms biofilm matures so high density and AIP is high so activates AgrC receptor and AgrA is phosphorylated so activates P3 and produces RNA III RNA III is an mRNA of hld (delta-hemolysin) and a regulatory RNA so represses adhesins and induces elements that drive invasion e.g. delta-hemolysin
102
Agr
accessory gene regulator operon with 2 promoters expresses AgrA/B/C/D D - AIP (autoinducing peptide) B - transmembrane, secretes mature AIP C - AIP receptor A - response protein activates P3 so RNA III
103
biofilms
structured clusters of cells enclosed in self-producing polymer matrix (from exopolysaccharides, proteins, nucleic acids), attached to surface and hides from environment like the immune system hard to desiccate, often anaerobic 80% of all microbial biomass is in biofilms
104
5 stages of biofilm formation
1. initial attachment - flagella and type I pili 2. irreversible attachment - LPS and type IV pili 3. maturation I - microcolonies form, produce sticky alginate and repress flagella so no movement 4. maturation II - quorum sensing 5. dispersion - release planktonic cells (single) from biofilm
105
P. aeruginosa biofilm
twitching motility used for maturation into microcolonies (stick to each other and sputum but not surface) and move along surface sigma factor 22 changes expression
106
Vibrio parahaemolyticus biofilm
switches between 2 flagella systems and interference with flagella leads to secondary swarming motility so flagella senses surface and interaction with it causes it to move along surface
107
synthetic cell biology
lecture 5

MCB - Molecular Bio (7 decks)

Brainscape helps you reach your goals faster, through stronger study habits.
© 2025 Bold Learning Solutions. Terms and Conditions