H2001 Final / Midterm 3 Flashcards
Which of the following questions would MD not be able to provide information about?
a) how blood cells interact in our body
b) membrane and embedded protein
c) protein binding site of drug
d) structural information for RNA
A
Describe the 2 heavy chains of myosin II
They wrap each other with 2 main parts:
N-terminal head domain: a globular motor domain that catalyzes ATP hydrolysis and interacts with actin
C-termini: arranged as extended alpha-helices wrapped around each other
Types of transport (2)
Passive transport: no E required
active transport: E required
See diagram on L26 slide 19 student verison
Actin
the “roads”; monomeric units (G-actin) come together to form long filaments (F-actin) that myosin walks along
Secondary transporters
pumps that use ion A’s gradient to move ion B
Cholesterol
steroid + alcohol; regulates membrane fluidity
Types of motor proteins (4)
1) Myosin
2) Actin
3) Kinesin
4) Dynein
Triacylglycerols
fatty acids are components of triacylglycerols
3 fatty acids esterified to a glycerol molecule (can be the same type of FA or different)
98% fatty acids in fats and oils
True or false: G proteins act as self-limiting switches in many processes
True
Which organelles have a double membrane in the endomembrane system? (3)
Nucleus
Mitochondrion
Chloroplasts
MAPK
mitogen activated protein kinase
Example of B-adrenergic pathway (steps 5 and 6 specifically) using glycogen
glycogen phosphorylase b-kinase becomes active when phosphorylated and stores glycogen in muscle and liver, anticipating energy needs signalled by epinephrine
How much energy do fatty acids provide, and what does this make it good for?
9kcal/g - great for storage
What is autophosphorylation?
Self phosphorylation
each ab monomer phosphorylates 3 critical Tyr residues near the carboxyl terminus of the B chain of its partner in the dimer
What is a good way to get a-linolenic
Fish products have a lot of EPA
Alpha-linoleic
omega-6, 18:2(delta9,12)
Pro-inflammatory effects
Structure of GPCRs
7 membrane spanning regions
C-terminal loop and tail has G-protein binding activity
3 families that bind ligands in different ways
What computational class won a Nobel Prize in 2013?
MD
Step 3 of the B-adrenergic pathway
activated G protein transmits signal to downstream effector protein, adenylyl cyclase
upon activation, dissociated beta and gamma subunits of the G protein form a By dimer, and the Gsa portion moves from the receptor to the adenylyl cyclase
The muscle fibers are composed of:
a) myofibrils
b) F-actin
c) thick filaments
d) all of the above
D
What are the 3 types of fatty acid signalling molecules?
Hormones, cofactors, binding sites
What type of myosin do we have in muscle?
Myosin II
What are the primary and secondary messengers in the B-adrenergic pathway?
Primary: epinephrine
secondary: cAMP
Step 3 of the mechanism of myosin motor
Phosphate gets released from myosin
Another conformational change occurs, myosin cleft closes
Myosin-actin binding is strengthened
What are the 3 components of the B-adrenergic pathway?
1) Epinephrine: produces signal
2) B-adrenergic receptor: receives signal
3) G proteins: acts as guanosine nucleotide exchange factor (GEF)
Example of gated ion channels
acetylcholine receptors on postsynaptic cells respond to binding of acetylcholine
How to use the alternate naming convention
omega refers to the position of the DBs as counted from the methyl end
a fatty acid would be “omega-3” if the location of the FIRST double bond is 3 Carbons counted from the methyl end
Types of mutations (3)
- point mutations (subs of one amino acid for another)
- deletions
- insertions
What are a couple of processes GPCRs are involved in? (5)
1) Sensory perception
2) Cell division
3) Growth and differentiation
4) Intracellular mvmts
5) Protein synthesis
What does Clostridium botulinium toxin do?
It is a protease; cleaves specific bonds in protein complex, preventing neurotransmission -> death
Adenylyl cyclase
integral membrane protein, with active site facing cytoplasm
What is MD not used for in biochemistry? (3)
- Understanding reactions
- Whole cells / organs
- long timescales
Termination Mechanisms:
Epinephrine concentration drops
Conc below Kd, epinephrine will dissociate, receptor returns to inactive form
Describe cholesterol role in terms of Lo / Ld
cholesterol makes Lo more fluid, Ld lipids less fluid
Insulin receptor signalling
Step 1
Insulin binds to alpha chains, activates Tyr kinase activity of B chains through autophosphorylation
What signal ignites the B-adrenergic pathway?
Epinephrine
What is the sarcomere in myofibril electron microscopy images?
A region from one Z-disk to the next
Short, medium, long chain fatty acids
Short: up to 8 Cs
Medium: 8 - 14 Cs
Long: more than 14 Cs
Tropomyosin and troponin
regulation of actin-myosin interaction
See myosin diagram L22 slide 13/14
In the Gibbs eqn for charged ions, what does C2 represent?
The concentration in the area the molecule is moving to
Signal transduction
how information (signal) is converted into chemical change (transduction) in living cells
See the two diagrams of myofibril electron microscopy images (L22 slides 23/24)
Monomer of actin
G-actin (375 residues)
How does cholesterol regulate fluidity (basic)?
High T - less flexible ring system / lower fluidity
Low T - more flexible ring system / higher fluidity
(clarify these, it might be the opposite?)
Liquid-ordered state (Lo)
low T; lipids in bilayer form a gel state, motion is restrained
What does a gap in sequences indicate?
sequence has been deleted or inserted
Stuff that happens at the ATP/ADP binding site can change the relative position of myosin’s pink and green domains, and modify the structure and actin binding. Why?
Protein marginal stability (barely fold). Weak and non-covalent bonds hold the protein together, and thus it is dynamic in conformation
Ion channels (more)
Acts as a controlled pore. When the channel is open, very rapid, specific mvmt of a large number of ions.
Pores can be ligand, voltage, or stress-gated
passive, not saturable
channel opens in a fraction of a ms, and only remains open for a few ms
Myosin
the motor protein; walks along actin filaments
Where do we see movement in our bodies (4)?
1) Cells
2) Organelles
3) Macromolecules
4) Us
Describe the B barrel structure of certain membrane proteins
20+ transmembrane segments (about 8 residues long) form B sheets that line a cylinder, stabilized by intrachain H bonds, alternating hydrophobic and hydrophilic residues
About how much shorter can the sarcomere become during contraction?
1/3 shorter
What is the sequence identity for the two sequences below:
S1: AGQWRKLTGH
S2: TGDWKRLTAH
50%
True or False: microfilaments composed of actin form one important part of the cytoskeleton and are key in muscle contraction
True
What organelles have a single membrane in the endomembrane system? (4)
Endoplasmic reticulum
Golgi apparatus
Lysosomes
vesicles
cAMP
cyclic adenosine monophosphate
i.e. ATP - 2 phosphate groups in cyclic form
Amplification in receptors
when enzymes active enzymes, the number of molecules increases geometrically in an enzyme cascade
Membrane rafts are characterized by the presence of:
a) sphingolipids and cholesterol
b) phospholipids and cholesterol
c) peripheral proteins
d) glycoproteins
A
What do we use when there are no DBs in the symbol naming?
n, -anoic acid
No delta needed, just write # carbons:0
Membrane lipids (3)
Glycolipids
Phospholipids
Cholesterol
Termination Mechanisms:
GTPase activator proteins (GAPs)
see diagram, L27 slide 17 students
Three types of lipid aggregates in water
1) Micelle: individual units are wedge shaped (Cross section of head greater than side chain)
2) Bilayer: individual units are cylindrical (cross section of head equals side chain)
3) Vesicle: forms spontaneously when bilayer sheet folds back on itself to form a hollow sphere (Aqueous cavity)
How do gated ion channels work? (3 main steps)
1) action potential arrives at terminal end of the pre-synaptic nerve cell, stimulating the release of vesicles containing acetylcholine
2) acetylcholine diffuses across synaptic cleft and binds onto target receptor
3) conformational change induced on channels upon binding, opening and exposing small and polar residues, allowing ions to pass through
see diagram on L25 slide 20
Step 1 of the mechanism of myosin motor
ATP binds to myosin
A cleft in the myosin molecule opens
Disruption of the actin-myosin interaction
Bound actin is released
Porins
proteins that allow certain polar solutes to cross the outer membrane of gram-negative bacteria; have B barrels lining the passage
Glycolipids
Sphingosine + fatty acid + sugar
(see diagram, L23 slide 27)
What are membranes good for? (4)
Separate intercellular from extracellular space
Boundaries
Role in signal transduction
gatekeepers (control passage of things in/out of cells)
Cell organelles also have ind membranes
Receptor tyrosine kinases:
a) bind two cAMP’s when active
b) do not regulate gene expression
c) are not prone to dimerization
d) autophosphorylate
D
Actin (more details)
One of the most abundant proteins
Highly conserved, greater than 95% sequence identity between species (as diverse as blue-green algae and humans)
Note: human skeletal muscle actin is identical in sequence to muscle actin in mouse, rabbit, rat, chicken, and cow
How do pumps work (general)?
pumps utilize E; transforms one form of E into another (ATPases pump using ATP - ATP hydrolyzed, conformation change, mvmt of ion/molecule)
Endocytosis vs. exocytosis
Endo: uptake into cell
Exo: excretion out of cell
both use vesicles
Describe the 2 classifications of anchored proteins
1) GPI-anchored: glycosylated derivatives of phosphatidylnositol; on C terminus; exclusively on outer phase of membrane
2) Palmitoyl: on cysteine residues; inside face of membrane
Benefits of computational modeling (3)
Safety (less risk)
Decrease time and cost of studies
Visualize processes
Insulin receptor signalling
Step 5: phosphorylation sequence
Raf-1, MEK, ERK are protein kinases that form a cascade, in which each kinase activates the next by phosphorylation
-Raf-1 phosphorylates MEK on 2 Ser residues
-MEK activated
-MEK phosphorylates ERK on a Thr and a Tyr reside
-ERK activated
How are myosin/actin arranged and how do they contract?
In filaments
Slide past each other to create contraction
True or False: membranes have complex compositions including lipids, proteins, and carbohydrates
True
Most extracellular ligands must interact with a membrane receptor; why?
Too large/hydrophilic to move through the membrane, so they act indirectly through receptors
F-actin
conglomerate of G-actin units (diameter of about 8nm)
Membrane channels
ion channels: allow ions to diffuse down concentration gradient; cause selective permeability to certain ions
General types of signal transducers (4)
- G Protein-coupled receptor: external ligand binding to receptor activates intracellular GTP-binding protein which regulates an enzyme, generating an intracellular secondary messenger
- Receptor enzyme (tyrosine kinase): explanation later
- Gated ion channel: channel opens or closes in response to concentration of signal ligand or membrane potential
- Nuclear receptor: hormone binding allows receptor to regulate expression of certain genes
What are the individual components of active cAMP-dependent PKA?
cAMP binds to R subunits, undergo a conformational change
Autoinhibitory domain of R moves out of the catalytic domain of C
R2C2 domain dissociates giving rise to 2 free, active C subunits
Active C subunits can now phosphorylate Ser/Thr residues on target proteins
How do solutes cross membranes? (3)
Simple diffusion if membrane is permeable enough
Membrane proteins: channels or transporters
membrane vesicles: large molecules or large amounts of small solutes
How are lipid bilayers formed?
They self assemble; the arrangement of hydrophobic portions with each other / hydrophilic with water favors higher system entropy
Signal transduction, the basics (5)
- A signal (ligand) interacts with the receptor
- activated receptor interacts with cellular machinery
- a second signal / protein activity change is produced
- metabolic activity changes (target cell)
- end of transuction event
What does each monomer of actin bind?
ATP - hydrolyzed into ADP
What is the common DB configuration in unsaturated FAs?
cis
iClicker for phylogenic tree
See Slide 24
About how many myofibrils does the muscle fiber contain?
1000
Sequence conservation
comparing the sequence of the same protein can give clues to the evolutionary relationship between species
True or false: membranes are very rigid and have fixed structures
False; they are very dynamic
Endomembranes
envelope organelles (endoplasmic reticulum, golgi apparatus, lysosomes, etc)
A charged ion going down the electrochemical gradient would cross the membrane through:
a) passive transport
b) active transport
c) facilitated diffusion
d) osmosis
e) it depends
C
What is the organization of skeletal muscle? (5 layers)
Skeletal Muscle <- Muscle fibers <- myofibrils <- thick and thin filaments <- myosin, actin (respectively)
Tropomyosin
binds to thin filament to prevent myosin from attaching to actin
Fatty acid suffixes
1: -enoic acid
2: -dienoic acid
3: -trienoic acid
based on #DBs
Co-transporters (2)
1) Symporters: 2 molecules transported in same direction
2) antiporters: molecules transported in different directions
(See ex slide 32 L25 of classroom)
What is true about peripheral proteins?
a) they are embedded in the bilayer
b) they are held by electrostatic interactions and H bonding
c) they are covalenlty bound to lipids
d) they are held by hydrophobic interaction
B
What does the Gs label represent?
G protein stimulates its effector
Review diagram of B-adrenergic pathway
L27 slide 11 students version
Describe the sodium-glucose symporter example in the intestines
2 Na ions and 1 glucose bind to symporter outside membrane. Conformational change; Na and glucose released in cell.
Specificity in receptors
signal molecule fits binding site on its complementary receptor, others don’t fit
Ligand gated ion channels
channel opens upon ligand binding; ions can bypass hydrophobic core of membrane
Describe the Na-K pump
1) cytoplasmic Na+ binds to the Na/K pump (3 sodium molecules bind) in cell
2) the pump is phosphorylated by ATP
3) the pump changes conformation, releasing Na+ out of cell
4) extracellular K+ binds to pump (2 molecules) leading to dephosphorylation
5) the pump returns to its original conformation
6) K+ released into cell
Steps of membrane fusion (4)
1) Membranes recognize each other
2) surfaces approach - removal of water molecules associated with lipid polar heads
3) local bilayer disruption, with further fusion of the outer leaflet of each membrane (hemifusion)
4) single continuous bilayer is formed
Step 4 of the mechanism of myosin motor
Release of phosphate causes the “power stroke” - conformation of myosin head returns to original resting state, its orientation relative to the bound actin changing to pull the tail of the myosin toward the Z disk
ADP is then released to complete the cycle
Phosphoglycerides
glycerol + 2 FAs + phosphate + alcohol
Gibbs Eqn for uncharged molecules
dG = RTln(c2 / c1)
where c1 is conc in the area the molecule is coming from (extracellular), c2 is concentration in the area it is moving to (cellular)
if dG > 0, molecule will not move spontaneously (active required)
if dG < 0, molecule will move spontaneously (passive)
What does GPCR stand for
G-Protein-Coupled Receptors
Molecular mechanism of myosin motor
A cycle with 4 major steps
Each cycle generates about 3 - 4pN of force and moves thick filament 5 - 10 nm relative to the thin filament
Peripheral proteins
electrostatic or H bond interactions with integral proteins and/or lipid head groups
Where does epinephrine bind?
Adrenergic receptors
Another method for naming fatty acids
Counts from the methyl end, typically only used for nutrition-related lipids
In which RTK domain does activation occur?
a) a monomers
b) b monomers
c) ligand binding site
d) membrane-spanning domain
B
Example of primary active transport
electrical signalling in neurons
ATP hydrolysis coupled to pumping (Na-K pumps), active because Na+ is way more concentrated outside cells
What state do long chain saturated fatty acids tend?
Lo
Variation in ligand types (4)
Small molecules (amino acids)
Gases (NO)
Soluble proteins (hormones, insulin)
Membrane anchored factors (cell antigens)
Insulin receptor signalling
Step 4
Activated Ras binds Raf-1, a protein kinase, activating it
Which of these is an example of a computational biochem method?
a) bioinformatics
b) MD
c) 3D structural prediction
d) AI
e) all of the above
E
Melting points of oils and fats
when the solid turns into liquid
Saturated: high melting point (butter, lard)
Unsaturated: low melting point (oils)
4 parts of the GTPase Switch
1) Gs with GDP bound is turned off; it cannot activate adenylyl cyclase
2) Contact of Gs with hormone-receptor complex causes displacement of bound GDP by GTP
3) Gs with GTP bound dissociates into alpha and beta-gamma subunits; Gsa GTP is turned on; it can activate cyclase
4) GTP bound to Gsa is hydrolyzed by the protein’s intrinsic GTPase; Gsa turns itself off. The inactive alpha subunit diffuses into the plane of the membrane and reassociates with the beta-gamma subunit
Types of adrenergic receptors (4)
Where are they found?
What do they do?
What are they?
alpha1, a2, beta1, b2
muscle, liver, adipose tissue
mediate changes in fuel metabolism
integral proteins with seven hydrophobic alpha-helices
Regulation of gene expression by insulin, 3 steps
1) insulin signal, on plasma membrane receptor
2) insulin-sensitive metabolic enzymes
3) nucleus, stimulates transcription of specific genes
What effect is responsible for the forming of the lipid bilayer?
Hydrophobic effect
What are the individual components of inactive cAMP-dependent PKA?
2 identical catalytic subunits (C)
2 identical regulatory subunits (R)
R2C2 domain
Autoinhibitory domain of each R subunit occupies substrate-binding cleft of each C subunit
See diagram L27 slide 13 students
G proteins
a member of the guanosine nucleotide binding protein family
function as on-off switches for intracellular signalling pathways
activate or inactivate ion channels
How do polar / charged ligands participate in signalling?
must bind on outside of cell, extracellular domains of cell-surface receptors
ex) peptide ligands (growth factors, insulin, neurotransmitters)
Membrane trafficking
exchange of material between 1 membrane and the next
Saturated vs. unsaturated fatty acids
Saturated: all single C-C bonds
Unsaturated: at least one double / triple C-C bond
If two genes or proteins have similar sequences, what can we suggest?
They have a common ancestor
More on channels
speed of transport
specificity
saturation
passive or active
allow transmembrane mvmt of ions at rates orders of magnitude greater than typical transporters (approaching limit of unhindered diffusion)
typically show some specificity for ion transport
not saturable with ion substrate
passive always
True or False: the “Mediterranean diet” has a ratio of omega-6 to omega-3 closer to 5:1, which is associated with lower risk of cardiovascular diseases
True
L25 slide 15 on classroom set
Protein logo
shows preferences in amino acid; larger occurances are more likely to be essential for protein function
Which sequence correctly identifies the order of events that take place during desensitization of the B-adrenergic receptor?
- Binding of B-arrestin to the receptor
- recruitment of Bark to the membrane by Gby subunits
- prolonged GPCR signalling leads to increases in free Gby subunits and transcription of BARK
- phosphorylation of Ser residues on the receptor
3, 2, 4, 1
When does simple diffusion happen?
Small, nonpolar molecules (passive)
G-protein types (2)
Monomeric type: involved in alteration of gene expression
Trimeric type (alpha, beta, gamma subunits): real G-proteins; 100 diff members with 1000s of receptors and functions; subunits can dissociate
Sphingolipids are a class of:
a) phospholipases
b) phospholipids
c) glycolipids
d) sterols
B
What does myosin do to walk along the filament?
Myosin binds F-actin and uses ATP hydrolysis to walk along the filament
What state do unsaturated fatty acids prefer
Ld
Review diagram of Steps of the mechanism of myosin motor
L22 slides 30 - 34
When does facilitated diffusion happen?
Large molecules / ions not able to bypass the hydrophobic core (passive)
Bioinformatics
compares sequence data, providing connections between species and changes over time
See L23 S24 (classroom notes) for an iClicker Q about naming fatty acids
E
Example of facilitated diffusion
Glucose transport into red blood cell (Specific; D-glucose but not L-glucose)
PUFA
polyunsaturated fatty acid
Termination mechanisms (2 parts, 3 steps (1) 2 steps (2))
Stimulus has ended:
1) epinephrine concentration drops below Kd for receptor
2) GTPase activity of G protein hydrolyzes bound GTP
3) cyclic nucleotide phosphodiesterase hydrolyces cAmp to 5’ AMP
Desensitization of receptor:
4) block G protein building sites on receptor (B-arrestin)
5) decrease number of receptors by triggering endocytosis of receptors (B-arrestin)
The myofibril part is composed of only thin filaments. Which part am I referring to?
a) A band
b) I band
c) Z disk
d) none of the above
B
Adrenergic
comes from adrenaline; fight or flight hormone
More on transporters
speed of transport
specificity
saturation
passive or active
catalyze transport at rates well below limits of free diffusion (slow)
bind substrate with high specificity
are saturable in the same sense as are enzymes
can be active or passive
What is the acetylcholine receptor?
Heteropentamer; 4 types of subunits (2alpha, beta, gamma, and delta chains)
integral membrane protein
acetylcholine binds each of 2 alpha subunits
What are the 2 major muscle proteins and what are they considered?
Myosin and actin; contractile force of muscle
Where does GTP/GDP binding happen in the G proteins?
Alpha subunit
How do small / hydrophobic ligands participate in signalling?
Cross membrane, bind to intracellular receptors in nucleus or cytoplasm
must bind to travel through bloodstream
ex: NO and steroid hormones, estradiol and testosterone, Vit D
What is the M line in myofibril electron microscopy images?
The center point of the A band, between two Z disks
Cell surface receptor 3 domains
- Extracellular ligand-binding domain
- Hydrophobic domain (spans across membrane)
- Intracellular domain: transmits a signal
Termination Mechanisms:
Cyclic nucleotide phosphodiesterase
reverses activation of PKA
L27 slide 18 students diagram
Integral proteins
embedded within the hydrophobic region of the bilayer
Sequence identity
fraction of positions that have exactly the same amino acid or nucleic acid (divide matching residues by total number)
What is skeletal muscle a form of?
Striated muscle
Hydrogenation of fatty acids
to improve shelf life / increase stability at high T, fatty acids can be prepared by partial hydrogenation
converts cis DBs to trans DBs
More about fatty acids
Hydrocarbons
Non-polar
12 - 20 Carbons
Even number of Carbons
Types of cell surface receptors (3)
Ligand-gated ion channels
G protein-coupled receptors
Receptor tyrosine kinases
MAPK family
phosphorylates Ser, Tyr, or Thr residues, depending on family and event
ERK, MEK, and Raf-1 in MAPK(K)(K) family
Membrane dynamics
membranes can change shape without losing integrity
phospholipids can change place
Insulin receptor signalling
Step 3: activation step
protein complex transfers the message from insulin receptor to end targets in the cytosol and nucleus
several intermediate proteins are involved:
-Grb2 (growth factor) has a SH2 domain
-SH2 domains binds to P-Tyr of IRS-1
-Grb2 also has an SH3 domain
-SH3 domain binds a proline-rich region in Sos
-Sos catalyzes replacement of GDP for GTP on Ras (G protein)
-> Ras activated
see cycle diagram L28 slide 12 students
What does trypsin do to myosin?
Breaks / cleaves the protein at the neck, producing light and heavy meromyosin chains
Secondary active transport
transport of a solute against its electrochemical gradient is coupled to transport of an ion down its electrochemical gradient so the overall free E change is negative (ATP hydrolysis required to maintain ion gradient)
ex) glucose transport in intestine (different from the mechanism in the red blood cells)
Gated ion channels
ion channels that respond to the binding of specific cellular event (ligand binding, voltage, stress)
How much of protein mass of muscle is myosin/actin?
80%
Receptor Tyrosine Kinases (RTKs)
basic def, extracellular vs. intracellular, end result
family of plasma membrane receptors with protein kinase activity
have an extracellular ligand binding domain and a cytoplasmic tyrosine (Tyr) kinase domain
intracellular Tyr kinase activity can then act through cascades
end result is to transduce signals about the metabolic state, including growth factors
See slides 21 - 24 on L26 for diagram practice
Myosin superfamily
they have a common ancestor; all move along actin filaments while hydrolyzing ATP.
What makes us move?
Molecular motors
Membrane pumps
Ion transporters: actively move ions against concentration gradient and create ion concentration gradients
ion binds to transporter and is transported across membrane
Consequences of mutations
If mutation is deletetion, it may cause disease or will be lost
if mutation is beneficial for function, it will be retained
Insulin receptor signalling
Steps 6 and 7: more phosphorylation and transcription
ERK enters nucleus, phosphorylates proteins such as Elk1
Elk1 modulates transcription of about 100 insulin-related genes
Glucose transport through GLUT1 is considered a passive transport because….
a) Glut1 contains a hydrophilic pore
b) an ion channel is used
c) a conformation change in Glut1 takes place
d) there is no ATP expenditure
D
When does primary active transport happen?
large molecules/ions moving against the electrochemical gradient
What are the 8 features of signal transduction and what are they? (in a few words)
- Specificity: ligand and receptor adapt to each other
- Sensitivity: high affinity
- Amplification: enzyme cascade
- Modularity: multiple domains, multiple enzymes, multiple functions
- Desensitization/Adaptation: overwhelming signals lead to desensitization
- Integration: multiple signals, one message
- Divergence: branching effect
- Localized response: confined regulation
What are some of the different classes of computational biochem (7)?
- Bioinformatics (function, sequence)
- Molecular modeling (function, structure)
- Molecular dynamics (structure, function)
- Structural prediction (Structure, sequence)
- homology modeling (Structure, sequence)
- Machine learning
- AI
Which signalling system actually moves a molecule across the plasma membrane, as opposed to only a conformational change in the receptor?
a) G protein-coupled receptors
b) nuclear receptors
c) gated ion channels
d) tyrosine kinase receptors
C
See diagrams of membrane fusion, L24 last couple slides
Receptor tyrosine kinases (general)
enzyme-linked receptors are located in cell surface; intracellular domains associated with an enzyme
Phospholipids
amphipathic
phosphoglycerides or sphingolipids
What is the form of information in cells?
Signals
Arrange these fatty acids from lowest to highest melting point
Lignocerate: 24:0
Linoleate: 18:2
Oleate: 18:1
Stearate: 18:0
Linoleate < Oleate < Stearate < Lignocerate
More on integral membrane proteins, describe:
a-helix sequence length
stabilization
potential for H bonding
a-helical sequence of 20-25 residues, long enough to span the bilayer
stabilitzed by intrachain hydrogen bonds and hydrophobic interactions
no backbone CO/NH groups inside the membrane (no potential for H bonds)
Functions of fatty acids (3)
Provide energy
Structural
Signaling
What is the chemical energy that is transformed into motion?
ATP
What does papain do when added to heavy meromyosin?
Separates the neck from the heads
Helps studying ATP hydrolysis
Paralogues vs. orthologues
paralogues: 2 homologues in same species
Orthologues: 2 homologues in different species
How can the ligand-gated ion channel be specific, using acetylcholine example
Acidic residues near end of each helix; anions cannot pass
Size of channel does not allow larger ions to pass
See diagram on L24 slide 12 (classroom)
True or False: GTP bound means inactive and GDP bound means active
False; opposite
What is the basic cycle for GPCR signalling in cholera?
Defective regulation of adenylyl cyclase (loses GTPase activity)
-> overproduction of cAMP
-> chronically active PKA
-> efflux of NaCl causes massive water loss
Where are omega-3s found?
Fish oil, flax, chia, safflower oil, olive oil
3 components of signal transduction through GPCRs
- A plasma membrane receptor with 7 transmembrane helical segments
- A G-protein that cycles btwn active (GTP bound) and inactive (GDP bound) forms
- Effector enzyme or ion channel in plasma membrane regulated by activated G protein
Hydropathy plot
shows regions of hydrophobic side chains, gives an idea which residues are on the outside or inside of the membrane
Desensitization: Block G protein sites on receptor
B-adrenergic receptor kinase (BARK) phosphorylates Ser residues near carboxyl terminus
G protein coupled receptor kinases, GRKs, play similar role
B-arrestin can now bind to the receptor, blocking interaction with the G protein
What factors impact melting point of oils/fats?
Higher chain length = higher melting point
more DBs = lower melting points
trans DBs = higher melting point
Desensitization: Decrease number of receptors
The arrestin receptor complex recruits proteins involved in vesicle formation, initiating membrane invagination and eventual sequestration of receptors in endosomes
What are GPCRs
large diverse family of receptors, respond to a variety of extracellular signals
Thick filaments
several hundred myosin molecules aggregate to form thick filaments.
Step 4 of the B-adrenergic pathway
When adenylyl cyclase couples with Gsa, it catalyzes the synthesis of cAMP from ATP
Concentration of cAMP, a second messenger, increases in the cytosol
Know general structure of phosphoglycerides; how do they connect? (See L23 slide 25)
FA to glycerol
Glycerol to phosphate
phosphate to alcohol
Why do we need a balance of a-linoleic and a-linolenic acid?
We need to regulate inflammation (need some but not too much)
MUFA
monounsaturated fatty acid
How do signals deliver information?
Captured by receptors and converted to cellular response (involves chemical process)
Homologous
have a common ancestor
20-30% identical amino acids
Review example of glucose transport
L25 slide 19 - 22 (students notes)
Computational biochemistry
what is it, what is it based on, what does it provide info on
experiment in computer (in silico modeling)
based on chemistry and physics and thermodynamics
provides info about molecular structure, chemical properties and non-covalent interactions
Insulin receptor signalling
Step 2: pathway on
receptor can now phosphorylate Tyr residues of other target proteins, such as insulin receptor substrate-1 (IRS-1)
upon phosphorylation, IRS-1 becomes a nucleation hub for a complex of proteins
What are membrane rafts good for?
organization of key proteins
membrane fusion (fusing 2 membranes together is critical for many cellular processes such as exocytosis, endocytosis, cell division, viral infection)
GPCR Signalling in Disease: Cholera
(activating vs. inactivating mutations, toxins, cholera toxin)
activating mutations in Ga lead to elevated [cAMP] (~40% adenomas)
inactivating mutations in Ga cause individuals to be unresponsive to hormones that act through cAMP
Toxins can interact with signalling mechanisms
Cholera toxin transfers an ADP-ribose to a residue of Gsa and blocks GTPase activity, causing massive water loss due to chronically active PKA, efflux of NaCl
B-adrenergic pathway
well-understood pathway that serves as a prototype for all GPCRs
What are the light / dark bands in myofibril electron microscopy images?
Light: I band
Dark: A band
When are neurotransmitters released in membrane fusion?
At synapses when intracellular vesicles loaded with neurotransmitter fuse with plasma membrane; SNAREs promote fusion
See the table about differences in composition of membranes
L24 slide 15 (classroom)
Alpha-linolenic
omega-3, 18:3(delta9,12,15)
anti-inflammatory effects
What are the helices named in ligand-gated ion channel?
M1, M2, M3, M4
M2 has polar and nonpolar side and thus lines the channel
Gibbs Eqn for Charged molecules
dG = RTln(c2 / c1) + ZFdeltapsi
where c1 is conc in the area the molecule is coming from (extracellular), c2 is concentration in the area it is moving to (cellular), Z is charge on ion, F is Faraday’s constant, deltapsi is transmembrane electric potential
The role of ATP hydrolysis in muscle contraction is…
a) break the interaction between actin and myosin
b) power up the re-attachment of myosin to actin
c) strengthen myosin-actin binding
d) a and b
e) b and c
B
Membrane rafts
region in membrane where sphingolipids associate with cholesterol; bilayer is thicker and less fluid
What is the role of protein kinase A in the B-adrenergic pathway?
a) it phosphorylates the Gsa for deactivation in a feedback inhibition loop
b) it deactivates adenylyl cyclase by phosphorylation
c) it phosphorylates specific Ser or Thr residues on target downstream enzymes
d) it mobilizes muscle and liver glycogen stores to provide the needed energy, as signalled by epinephrine
C
Essential fatty acids
body can’t produce it, so need it from diet
a-linolenic acid (eicosapentaenoic acid, EPA)
a-linoleic acid (arachidonic acid)
Symbol Naming fatty acids
chain length and number of double bonds, separated by a colon
Numbering begins at the carboxyl carbon
positions of double bonds are indicated by a delta and superscript number
Use suffix
The Insulin Receptor (structure)
2 identical alpha chains, outer face of membrane (insulin binding sites)
2 transmembrane B subunits, facing the cytosol, with protein kinase activity
Phosphate from ATP transferred to OH of Tyr residues on target proteins
What do channels do?
Allow ions to diffuse without ever interacting with the membranes hydrophobic core
How much do cells spend of the ATP they produce?
30-70%
Step 2 of the mechanism of myosin motor
ATP is hydrolyzed
Protein changes to a high-energy conformation
Myosin head moves and changes orientation in relation to actin thin filament
Myosin binds weakly to F-actin subunit closer to the Z disk than the one just released
What does your body need for a fight or flight reaction?
Energy
Glucose transport using facilitated diffusion
1) glucose in blood plasma binds to transporter and lowers activation energy
2) conformational change from T1 to T2 conformation, influencing the transmembrane passage of the glucose
3) glucose is released into the cytoplasm
4) transporter returns to T1 conformation
Steps 5 and 6 of the B-adrenergic pathway
cAMP allosterically activates PKA (protein kinase)
PKA catalyzes phosphorylation of specific Ser and Thr residues of targeted proteins
Phylogenic trees
lengths of lines proportional to number of amino acid substitutions
shorter lines = closer evolutionary relationship
Structure of thick filaments
Tails pack end to end in a staggered array, allowing globular heads to project from the sides of the molecule
What type of information do animal cells exchange?
- Concentration of ions / glucose in extracellular fluids
- interdependent metabolic activities in tissues
- correct placement of cells during development
North American omega-6 to omega-3 ratio
20:1; associated with higher risk of cardiovascular disease
What state do short chain saturated fatty acids tend
Ld
Nerve impulse
release of Ca2+ from sarcoplasmic reticulum
Ca2+ binds to troponin
conformational change in tropomyosin-troponin complex, myosin can bind, and contraction happens
Receptor proteins (location, binding, mediation)
may be in membrane, other organelles, or cytosol
specific binding of ligands, inducing conformational changes. Have effector specificity
may mediate a variety of actions (i.e. cell - cell signalling, adhesion, endocytosis, signalling cascades, gene regulation)
Steps 1 and 2 of the B-adrenergic pathway
Epinephrine binds the B-adrenergic receptor
receptor undergoes allosteric transitions
facilitates displacement of GDP and binding of GTP, activating the G protein
How does RTK activation happen?
Autophosphorylation
each B subunit phosphorylates 3 essential Tyr residues near C-terminus of the other B subunit, results in opening of the active site
What bands are produced in myofibrils when viewed using electron miscroscopy (3)?
1) I band: only thin filaments (actin)
2) A band: thick filaments + region where tick and thin filaments overlap (actin and myosin)
3) Z disk: where thin filaments attach
Lipid building blocks
Fatty acids
Check the broader def of 8 features L26 slide 7 (student notes)
MAPK cascades
mediate signalling initiated by a variety of growth factors
amplify the signal by many orders of magnitude
Where does GTPase come from?
Gsa has internal GTPase activity that converts bound GTP to GDP, inactivating Gsa
Molecular dynamics
tells us how atoms interact over time
based on classical mechanical physics
True or False: during contraction, thick and thin filaments slide past each other and Z-disks get farther away
False; they get closer
Sphingolipids
sphingosine + 1 FA
Sphingosine: amino alcohol + long unsaturated hydrocarbon chain
What do motor proteins do (basic)?
Transform E into movement
Thin filament structure
F-actin + troponin + tropomyosin
Which feature of signal transduction is defined by a primary signal is intensified by orders of magnitude?
Amplification
Conserved Structure/Function relationship in myosin
tail domains of different myosins differ in sequence significantly more than head domains
this is so myosins may interact with a larger number of cargoes with their tails — the goal in each case is the same, to walk along the actin filaments, which is why the motor / heads are similar
Embedded or integral proteins
hydrophobic portions interacting with fatty acyl chains of lipids
Basic structure of myosin II
4 light chains
2 heavy chains
Structural prediction
tells us structure of protein, nucleic acid, or carb
Epinephrine cascade; what does it do, what does it need, how long does it take?
amplifies the hormonal signal by orders of magnitude
only need a low conc of epinephrine for activity
signal leads to intracellular changes within fractions of a second
How does cholesterol increase / decrease rigidity?
Increase: interacts with unsaturated fatty acids, compacting them and constraining their motion (at high T we want to increase rigidity)
Decrease: associates with saturated fatty acids, making the bilayer more fluid (we want the membrane less rigit/more fluid at low T)
What are membrane rafts stabilized by?
Hydrophobic effect
Liquid disordered state (Ld)
high T; lipids in a bilayer have lots of motions, i.e. rotations, lateral diffusion, acyl chain mvmt
Ras/MAP kinase signal transduction
MAPK
Membrane fusion
Biological membranes can fuse with each other
Describe a fatty acid given by the symbol name 18:1(delta^9)
Has 18 Carbons, 1 double bond starting at C #9, counted from the carboxyl C
Sequence of events in RTKs (4)
- Ligand binding: specific recognition and interaction
- Dimerization: two ligand bound receptors come together
- Receptor activation: each receptor activated by autophosphorylation
- Signal transduction: active dimer now able to phosphorylate other proteins
Describe the 4 light chains of mysoin II
they wrap around the neck of each heavy domain and provide stiffening in the neck domains and in some cases have a regulatory role
Allosteric
binding at 1 site effects binding at another site, using conformational changes
What do thin filaments do?
Regulate muscle contraction so it only occurs following a nerve signal (see diagram, L22 slide 19)
Anchored proteins
covalently linked to one or more lipids (ex GPI)
Why do molecules move from higher to lower concentration?
Entropy - energy distribution over a larger area
Trans fats
raise LDL (bad cholesterol), decrease HDL (good cholesterol), and have potential adverse health consequences
