Midterm 2 Review Flashcards
1
Q
What is Illumina sequencing
A
Visualization of DNA sequencing.
Step 1 involes DNA Polymerase + Flouroscent reveraable Terminator NTPs,
Step 2 Flourcent tag and terminator removed from nt
2
Q
How many times is Illumina sequencing repeated
A
10^6
3
Q
how can one obtain snapshot of gene expression?
A
analysis of mRNA
4
Q
What is a RNA-Seq
A
- quantitative analysis of transcriptome
- globally sequence all the RNAs produced
by a cell - complete collection of RNA produced
by a cell under a certain set of conditions - relative abundance of a particular sequence
- # of copies present
5
Q
What is a single-cell RNA seq
A
adapted to catalog the mRNAs produced
by single cell
* single-cell RNA seq
* the mRNA from each cell are converted
into cDNA, each tag with a “barcode”
* can identify genes that
are coordinately regulated
* can be used to analyze the
changes in gene expression
6
Q
What is In-situ hybridization
A
- allows a specific NA sequence to be visualized
in its normal location
7
Q
What is Ribosome Profiling
A
which mRNA are being actively translated
to produce proteins
* extract of a cell or tissue
is exposed to ribonuclease
(RNase)
* protected RNA segments
are released from the
ribosomes, converted to
cDNA and sequenced
* mRNA sequences covered
by bound ribosomes are
spared from digestion
8
Q
What is the best way top study the function of a gene
A
to see what happens to an organism
when the gene is inactivated
9
Q
What is RNA interference
A
introduction of dsRNA molecule into cell or an
organism that matches the gene sequence to be
inactivated
dsRNA is cleaved and processed
by special RNAi machinery to
produce shorter, ds fragments
called small interfering RNAs
(siRNAs)
* siRNA → ssRNA fragments
hybridize with target gene mRNAs
and direct their degradation
Drosophila,
C. elegans
10
Q
What is Targeted gene replacement
A
in vitro
* change/alter regulatory region of gene
* introduce changes to coding sequence
* reintroduce back into organism to determine
gene function
* mutant organism transgenic organism
* introduced gene is called transgene
11
Q
How is Targeted gene replacement done in mice
A
ES Cells grow in culture.
Introduce a dna fragment into cells containing altered gene.
Take cells from prolifered colony and inject into early embryo
12
Q
What is an example of Targeted gene replacement done in mice
A
Leptin deficient
13
Q
What is A conditional knockout mice
A
disrupt a known gene more selectively
conditional knockouts are useful if the gene
under study is critical during embryonic
development
14
Q
What is the CRISPR System
A
powerful new gene editing method
method to turn
selected genes
on and off
15
Q
What is the key protein in the CRISPR System
A
Cas9. Used to break double strand
16
Q
What happens when catalytically inactive CAS9 fused with transcription activator
A
Target gene is on
17
Q
What happens when catalytically inactive CAS9 fused with transcription repressor
A
The target gene is off
18
Q
What is DNA Cloning
A
allows production of recombinant protein
in large quantities
efficient production is usually accomplished
using specially designed expression vectors
different expression vectors are designed for
use in bacteria, yeast, insect or mammalian cells
19
Q
Vectors/plasmids
A
circular recombinant dsDNA
modified bacterial plasmids
each plasmid contains
* replication of origin
* allows production
of large amount
within bacterial cells
* expression vectors transformation
* selection marker
20
Q
How are DNA fragments inserted into a plasmid
A
EcoR1 cleaves circular DNA. DNa fragment is covalently linked and sealed with DNA ligase
21
Q
How does Haelll cur DNA
A
Cuts at a sequence of 4 nt pair down the middle, leaving 2 blunt ends
22
Q
how often does Haelll cut
A
occur every 256 nt
(1 in 44) purely by chance
23
Q
how often does a restriction ezyme be expected to cleave
A
occur every 256 nt
(1 in 44) purely by chance
24
Q
What are some of the functions of proteins
A
- signal proteins
- motor proteins
- structural proteins
- transport proteins
- storage proteins
- enzymes
- transporters and
channels - receptor proteins
- specialized proteins
25
What is the most chemically know complex structure
Proteins
26
What type of bonds hold together AA
covalent peptide bonds
27
What side of the Protein is positive
N terminus
28
What end of a protein is negative
C terminus
29
How are peptide chains formed
by the removal of water
30
What are the 5 parts of a protein
Central carbon, Hydrogen, Carboxyl group, amnio group, R chain
31
What is the PH of a protein
7
32
Where is the peptide bond on a protein formed
Removal of OH on carboxly group and removal of H on amino group. C bonds to N. Amide Linkage
33
What type of bonds help fold proteins
Hydrogen Bonds,
Van der walls
Electrostatic Atrraction
Hydrophobic force
34
What is a hydrophobic force
has a central role in determining shape of the protein. Polar side chains on the outside of protein, where non polar sidechains are on the inside core
35
WHat are Chaperons
used to asssist protein folding
36
What is the most structurally diverse macro moleucule
proteins
37
What is the average length of a protein
50 - 2,000 aa long
38
What are 2 regular protein folding patterns
a-helix , b-sheet
result from hydrogen bonds that form between
the N-H and C=O groups in the polypeptide
backbone
39
What is a coiled-coil protein shape
* 2-3 a helices
wrap around
to form a stable
structure
* a keratin
40
What is a B Sheet
* antiparallel or parallel
* by convention;
* arrows point toward
the C-terminus
* produce a very rigid,
pleated structure
41
WHat is a subunit
each polypeptide chain
42
What is a protein binding site
* any region on a protein surface that interacts
with another molecule through set of
noncovalent bonds
43
What is a dimer
* two polypeptide chains form a symmetrical
complex of protein subunits
* held together by
interactions between
two binding sites
44
What are the Major filaments of the cytoskeleton
actin
tubulin
* intermediate filaments
* assembly into rope-like structures
* gives cytoskeleton mechanical strength
45
What is a ECM protein
Extracellular matrix protein
46
What is the purpose of a ECM protein
help bind cells together to form tissues
* secreted by cells into their surrounding
* often assembled into sheets or long fibrils
47
What is Collagen
* long, regular
triple helix
ECM Protein
48
What is elastin
* rubberlike elastic
meshwork
* elastic fibers
ECM Protein
49
How are ECM Proteins Stabilized
covalent disulfide bonds
50
What is a covalent disulfide bonds
Bonding of Sh-Sh or S-S
51
What is a Interchain disulfide bond
S-S between 2 protiens
52
What is a Intrachain disulfide Bond
S-S bond within same protein
53
What is a ligand
any ion, small organic
molecule or a macromolecule bonded to a protein. Non covelent bond to protein
54
What is an enzyme
chemically transform the ligands to which
they bind
* ligand for an enzyme is called a substrate
* substrate is converted into chemically
modified product
55
How can we determine enzymes performance
Michaelis constant, KM
56
What is a Lysozyme
* catalyzes hydrolysis
* adds a molecule of water to a single glycosidic
bond between two adjacent sugar groups in the
polysaccharide chain
57
What is a polysaccharide chain
* an enzyme that cuts polypeptide chains
* tightly bound zinc ion in its active site
* zinc ion forms a transient bond
with one of the substrate atoms
assisting the hydrolysis reaction
during the cleavage of a peptide
bond
58
What is a Biotin
* found in enzymes that transfer a
carboxyl group from one molecule
to another
* biotin forms a covalent bond to the –COO-
group to be transferred, thereby producing
an activated carrier
59
What is feedback inhibition
* an enzyme acting early in a reaction pathway
is inhibited by a molecule produced later in
that pathway
60
What type of regulation does feedback inhibiton create
negative regulation
61
What is an example of feedback inhibition
bacterial biosynthetic pathway
* aa controls the 1st enzyme
specific to its own synthesis
62
What does AlloSteric mean
* allo “other”
* stere “solid” or “shape”
63
What is an allosteric protein
* many enzymes contain at least two different
binding sites
* active site
* regulatory molecules binding site
64
What is Reversible Phosphorylation
is a common mechanism for regulating
protein activity
Kinase turns on protein while Phosphatase removes P and turns off protein
65
What are the 3 Reversible Phosphorylation
Ser, Thr, Tyr
66
What are 3 examples of Covalent modifications on proteins
* histones – acetylation/methylation
* lipid-linked proteins
* Ub-targeting for degradation
67
What is p53
* transcription regulator
* regulates cell’s respond to DNA damage
* covalently modified on at least 20 sites
68
Regulatory GTP-binding proteins are
switched on and off by the gain and loss
of a P group
* active protein can shut itself off by hydrolyzing
its bound GTP → GDP + phosphate
* P is not enzymatically transferred from ATP
* P is part of guanosine triphosphate (GTP)
69
Motor proteins
help to move chromosomes to opposite
ends of the cell during mitosis
* move organelles and macromolecules
along cytoskeletal tracks
* can move by undergoing a series
of conformational changes
* reversible changes
* random wandering
70
What allows Motor proteins to produce direct movement
* conformational changes must be unidirectional
* one of the steps must be irreversible
* coupling one of the conformational changes
to the hydrolysis of an ATP molecule
* motor proteins are also ATPases
71
how do proteins locate their interacting partners
and assemble into complexes that are activated
only when and where they are needed?
scaffold proteins
72
What is a scaffold proteins
* large molecules that contain binding sites
recognized by multiple proteins
* can greatly enhance a rate of particular
chemical reaction or cell process
73
Protein purification steps
standard approach to purify protein
* chromatography steps
* use different materials to separate individual
components of a complex mixture into
fractions based on protein properties
74
What are antibodies
immunoglobulin proteins produced by the
immune system in response to foreign
molecules
* each Ab binds to a particular target molecule,
an antigen, with remarkable specificity
75
What is the shape of an antibody
are Y-shaped molecules with two identical
antigen-binding site
2 light chains and 2 heavy chains
76
How are antibody binding sites formed
formed from several
loops of polypeptide chain that protrude from
the ends of a pair of juxtaposed protein
domains
an enormous diversity of antigen-binding sites
can be generated by changing the length of aa
sequence of these “hypervariable loops
76
how are anti bodys specified to antigens
* each Ab made by an
organism has a distinct
antigen binding site
* each Ab recognizes
an antigen with
great specificity
77
What produceds antibodies
B lymphocytes
B cell is stimulated
to proliferate
and to make
and secrete
more of the
same Ab
78
How can antibodies be used to purify molecules
Immunoprecipitation
79
How can antibodies be used as molecular tags
Microscopic detection
* Biochemical detection
80
1st protein sequence
1955 insulin
81
What is mass spectrometry
his technique determines the exact mass
of every peptide fragment in a protein
and/or mixture of proteins
82
What are 3 ways proteins 3D structures determined experimentally
* x-ray crystallography
* NMR spectroscopy
* cryo-electron microscopy
83
3D structure:
X-ray crystallography
uses x-rays, which have a diameter of
a hydrogen atom
* probe the structure of proteins at an atomic
level
84
NMR spectroscopy
if protein is less than 50 kDa
* for many atoms, hydrogen in particular,
the nucleus is intrinsically magnetic
* solution of
pure protein
85
3D structure:
cryo-electron microscopy (cryoEM)
* frozen sample examined
by transmission electron
microscopy
* algorithms sort
particles into sets
86
What is a cell membrane
act as selective barriers
87
What are Biological membranes made up of
* lipids
* proteins
88
What functions is the plasma membrane involed in
receiving information
import and export of molecules
movement and expansion
89
What are the amphiphilic properties of the lipid bilayer
Polar head Hydrophilic
Non polar tail is hydrophobic
90
What are the 3 different types of membrane lipids
* phospholipid
* glycolipid
* sterol
91
What is an example of a Purely hydrophobic lipid molecules
triacylglycerol
* main constituent of animal fats
* function as energy
reservoirs in animal
cells
92
What is unique about phospholipid
contains serine
93
What is unique about glycolipid
contains GAL
94
What is unique about sterol
1 hydrocarbon tail
95
What is unique about triacylglycerol
3 hydrocarbon tails
96
What is unique about a lipid bilayer
* lipid bilayer is flexible – able to bend
* membrane vesicle ~ 25nm in diameter
97
How does fluidity of lipid bilayer composition change
depends on
* phospholipid composition
* nature of the hydrocarbon tails
* two major properties of hydrocarbon tails
affect how tightly they pack into bilayer
* the length
* the # of double bonds
98
How can hydrocarbon tails vary
* vary in length between 14 and 24 carbon atoms
* 18-20 atoms most common
* shorter chain length increase the fluidity of the bilayer
* typically,
* one chain contains only single bonds
between carbons saturated
* the other chain includes one or more
carbon-carbon double bond unsaturated
99
How is fluidity in animal cells moderated
* inclusion of cholesterol
* constitutes ~20% of plasma membrane lipids
* cholesterol can fill the spaces between adjacent
phospholipid molecules left by the kinks in their
unsaturated hydrocarbon tails
* less flexible
* less permeable
100
What is the importance of membrance fluidity
* allows protein-protein interactions
* cell signalling
101
Where are Glycolipids found
* mainly in PM
* only in the noncytosolic half of the
lipid bilayer
101
How are new phospholipid manufactured
free fatty acids as substrates by enzymes
bound to the cytosolic surface of the ER
* newly synthesized
PLs are deposited
onto cytosolic half
of the bilayer
102
Where are glycolipids formed
assembled in Golgi
102
Diffusion across artificail bilayer- small nonpolar molecules
O2 CO2 N2
major goes across while some doesnt
103
Diffusion across artificail bilayer- Uncharged polar molecules
H20, ethanol, glycerol
little amount goes across
103
Diffusion across artificail bilayer- Ions
No diffusion
104
Diffusion across artificail bilayer- Large uncharged polar molecules
Glucose, amino acids
little defussed,most doesnt`
105
What are membrane proteins
in animals, proteins constitute ~50%
of the mass of PM
* a cell membrane typically contains
about 50 times the number of lipid
molecules compared to protein molecules
105
what is transmembrane (TM) proteins
* amphiphilic proteins
* hydrophobic region
* hydrophilic regions
* in the interior of the bilayer
* exposed to aq environment
on either side of the membrane
106
What is a monolayer associated protein
* located almost entirely in the cytosol
* associated with the cytosolic half
of the lipid bilayer by an amphipathic
helix exposed on a surface of the
protein
106
What is a lipid linked protein
* proteins lie entirely outside the bilayer
and are attached to the membrane by
one or more covalently attached lipid
groups
106
What is a protein attached protein
these proteins are held in place by
interaction with other membrane
proteins
106
What is a transmembreane a helix
* composed of aa with hydrophobic side chains
* interact with the hydrophobic
tails of the lipid molecules
* polypeptide backbone itself is hydrophilic
as peptide bonds are normally polar
* atoms of the hydrophilic
polypeptide backbone
form hydrogen bonds
with one another
107
What is an example of a single-pass
transmembrane
proteins
receptors
108
What is an example of a multipass
transmembrane
proteins
Channels
109
What is a A transmembrane sheets
beta sheets arranged into barrel structure
allow passage of small
nutrients, metabolites
and inorganic ions
109
What is Epithelial cells
* asymmetric distribution of membrane proteins
* maintained by barrier formed by tight junction
109
What is a Membrane Transport Proteins
span the lipid bilayer and provide private
passage across the membrane for
selected substances
110
What is facilitated transport
* passage across cell membrane must be
accelerated by specialized membrane
transport proteins
* multipass TM proteins
111
What is a transporter
* shift small organic molecules or inorganic
ions from one side of the membrane to the
other by changing shape
111
What is a channel
* form tiny hydrophilic pores across the
membranes through which substances
can pass by diffusion
* ion channels
112
What is the intracellular concentration of na
5-15
113
What is the intracellular concentration of K
140
113
What is the intracellular concentration of mg
0.5
113
What is the intracellular concentration of Ca
10^-4
113
What is the intracellular concentration of H
7 x10^-5
114
What is the intracellular concentration of CL
5-15
114
What is the extracellular concentration of Na
145
115
What is the extracellular concentration of K
5
115
What is the extracellular concentration of Mg
1-2
116
What is the extracellular concentration of Ca
1-2
116
What is the extracellular concentration of H
4 x 10^-5
117
What is the extracellular concentration of Cl
110
117
What is membrance potential
electrical charges inside and outside of the
cell are generally kept in balance, however,
tiny excess of +ve or –ve charge, do occur
* organism and cell type: -20 to -200 millivolts
117
What is passive transport
no expenditure of energy by the
membrane transport protein
* all channels and many transporters
act as conduits
for such passive
transport
high to low concentration gradient
117
What is active transport
* movement of a solute against concentration
gradient
* requires an input of energy
* carried out by special transporters
* pumps
* hydrolysis of ATP
117
what is the electrochemical gradient
* membrane potential and concentration gradient
govern the movement of uncharged molecules
and charged solutes across a cell membrane
118
What Aquaporins
* specialized channels to facilitate the flow
of water
119
what is osmolarity
total concentration of solute particles
inside of the cell
* generally exceeds the [solute]
outside of the cell
120
What are the 3 Conformational states of the glucose passive transpot proteins
outward-open
state
* occluded
state
* inward-open
state
120
What are the 3 types of active transport
Gradient driven pumps
ATP driven pumps
Light driven pumps
120
What is a NaK Pump
transports Na+ out of the cell as it carries K+ in
* uses energy derived from ATP
* accounts for 30% or more total ATP consumption
120
What is the cycle length in time of the NAK pump
10 miliseconds
120
What is the CA pump
moves Ca across membrane
120
What is Symport
Coupled transport of molecules that move in the same direction
121
What is Antiport
Coupled transport that moves in opposite direction
121
What is uniport
Transport of 1 intem in a direction. passive transport, not a pump
122
What is a Glucose-Na symport pump
* cooperative binding of Na+ and glucose
* uses the electrochemical Na+ gradient
to drive the active import of glucose
122
What is the primary function of membrane transport proteins?
B) To selectively transfer substances across membranes
122
Which type of transport protein forms tiny hydrophilic pores in the membrane?
123
The movement of solutes against their concentration gradient requires:
Active transport
123
What is the typical range of resting membrane potential in cells?
-20 to -200 millivolts
124
The Na+-K+ pump moves
3 Na+ out, 2 K+ in
125
In the Glucose-Na+ symport system, glucose is
Transported actively using the Na+ gradient
126
What ion do plant, fungal, and bacterial cells primarily use for active transport instead of Na+?
H+
126
What is the function of the ATP-dependent H+ pump in lysosomes?
To maintain an acidic environment
126
Phospholipids are considered amphiphilic because:
They have both hydrophilic and hydrophobic regions
126
The fluidity of the lipid bilayer depends on:
The length and number of double bonds in hydrocarbon tails
126
How does cholesterol influence membrane fluidity in animal cells
Prevents phospholipids from packing too closely
127
Which type of membrane protein spans the entire lipid bilayer?
Transmembrane proteins
128
A β-barrel transmembrane protein is typically found in
The mitochondria and bacterial outer membranes
129
Scramblase plays a role in
Randomly distributing phospholipids between bilayer halve
129
Flippase is responsible for
Flipping specific phospholipids using ATP
129
Tight junctions in epithelial cells
Prevent lateral diffusion of proteins
130
The glycocalyx (carbohydrate layer) of the plasma membrane is important for
Cellular protection and recognition
130
FRAP (Fluorescence Recovery After Photobleaching) is used to study
Protein movement in membranes
130
Detergents are used in membrane protein studies to
B) Dissolve proteins and remove them from membranes
131
Which type of lipid is only found in the non-cytosolic side of the membrane
glycoolipids
131
What structure allows some bacteria and mitochondria to transport small molecules across their outer membrane?
Porins (β-barrel proteins)
132
The fluid mosaic model of membranes suggests:
A) Membranes are solid structures
B) Proteins are fixed in place
C) Lipids and proteins move laterally
D) Carbohydrates are embedded inside the membrane
C) Lipids and proteins move laterally
132
Which transport method does NOT require ATP?
A) Na+/K+ pump
B) Facilitated diffusion
C) Endocytosis
D) Primary active transport
B) Facilitated diffusion
132
Why do plant and fungal cells use H+ pumps instead of Na+ pumps?
A) They lack ion channels
B) Their extracellular environment lacks Na+
C) They use proton gradients to drive solute transport
D) They do not use active transport
C) They use proton gradients to drive solute transport
133
Which of the following is NOT a function of membrane proteins?
A) Signal transduction
B) DNA replication
C) Transport of molecules
D) Cell adhesion
B) DNA replication
134
Which scenario would cause a cell to swell?
A) Placing it in a hypertonic solution
B) Placing it in a hypotonic solution
C) Increasing Na+ outside the cell
D) Blocking all ion transport
B) Placing it in a hypotonic solution
134
What would happen to membrane fluidity if cholesterol was removed?
A) The membrane would become too rigid
B) The membrane would become too fluid and permeable
C) The membrane would become hydrophobic
D) The membrane would break apart
The membrane would become too fluid and permeable
135
What is the primary advantage of next-generation sequencing (NGS)?
A) It is more expensive than traditional sequencing
B) It allows rapid sequencing of entire genomes
C) It only works on bacterial genomes
D) It does not require DNA amplification
B) It allows rapid sequencing of entire genomes
136
What does RNA-Seq analyze?
A) DNA replication rates
B) Protein structures
C) Gene mutations
D) mRNA expression levels
D) mRNA expression levels
136
What is a key feature of single-cell RNA sequencing?
A) Uses gel electrophoresis for visualization
B) Each mRNA is tagged with a unique barcode
C) It can only be used on bacteria
D) It does not require cDNA synthesis
B) Each mRNA is tagged with a unique barcode
137
How does RNA interference (RNAi) work?
B) It degrades mRNA using siRNA
137
What is a transgene?
A) A foreign gene introduced into an organism
B) A gene with a missense mutation
C) A gene that has been deleted from the genome
D) A gene that only functions in bacteria
A) A foreign gene introduced into an organism
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