week 6 - 8 Flashcards
The barrier to nuclear entry or
exit
Nucleus is surrounded by a
double membrane
– Continuous with the
rough endoplasmic
reticulum (ER)
Proteins destined to the nucleus are:
- Translated on free ribosomes in the cytoplasm
- Bound by “carrier” proteins
- Transferred through pores in the nuclear
membrane (nuclear pore complexes) - Localized in the nucleus upon recognition of
protein binding partners
Structure of the nuclear pore complex
NPC is one of the largest protein
assemblies in the cell
- NPC is embedded across the
double membrane
Ions, small metabolites and
proteins less than about 40
kDa can diffuse passively
through the pore
– Diameter for passive
diffusion 9 nm
* Large proteins and
ribonucleoprotein
complexes cannot just
diffuse passively
– These are actively
transported through the
pore
The nuclear localization sequence
- experiment
Proteins synthesized
in the cytoplasm can
be imported to the
nucleus if tagged with
a nuclear localization
signal (NLS)
The nuclear localization signal (NLS)
* Proteins destined for the nucleus contain one or more NLS
* Examples of NLS
– Monopartite: PKKKRKV
– Bipartite: KRX(10-12)KKKK
Note the high proportion of basic amino acid residues Lysine (K)
and Arginine (R)
Nuclear Import Summary
Nuclear import requires importins to transport “cargo” through nuclear pores.
Cargo Binding: Importin binds the cargo and interacts with nucleoporins, which have Phe-Gly repeats.
Cargo Release: Inside the nucleus, RAN-GTP binds to importin, causing a conformational change and release of the cargo.
Cycle Reset: The importin-RAN-GTP complex exits the nucleus to restart the process.
Import is uni-directional due to:
Importin diffusion driven by a concentration gradient.
Energy from GTP hydrolysis maintaining the system’s imbalance.
Recycling Ran Summary
Inside the nucleus, Ran-GEF facilitates the exchange of GDP for GTP, converting Ran to its active form (Ran-GTP).
In the cytoplasm, Ran-GAP activates Ran’s GTPase activity, hydrolyzing GTP to GDP, resetting Ran for another nuclear import/export cycle.
Protein Recognition for Export Summary
Proteins destined for export from the nucleus have a leucine-rich nuclear export signal (NES).
The NES follows a consensus sequence:
Φ X₁₋₃ Φ X₂₋₄ Φ X Φ,
where:
Φ = hydrophobic residues (L, I, F, V, M).
X = any amino acid.
Ran-Dependent Nuclear Export Summary
Complex Formation: Cargo with a nuclear export signal binds to exportin and Ran-GTP, forming a triple complex.
Cargo Release: In the cytoplasm, Ran-GAP converts Ran-GTP to Ran-GDP, causing a conformational change that releases the cargo.
Cycle Reset: Exportin, along with Ran-GDP, re-enters the nucleus to restart the process.
Ran-independent mechanism for
nuclear export of mRNAs
- Nucleoplasm –
- Heterodimeric NXF1/NXT1
nuclear export receptor
complex binds to mRNA-
protein complexes (mRNPs). - Complex diffuses through
NPC by transiently interacting
with FG nucleoporins. - Cytoplasm – An RNA helicase
(Dbp5) located on the cytoplasmic
side of the NPC uses ATP energy
to remove NXF1 and NXT1 from
the mRNA. - Recycling system – The Ran-
dependent import process
recycles free NXF1 and NXT1
proteins back into the nucleus.
The nuclear pore complex allows:
1. Passive diffusion of smaller molecules
2. Import of proteins
3. Active transport of very large molecules
4. All the other options
all the options
Which proteins participates in nuclear export of mRNA?
1. ribosome
2. exportin
3. NXF1/NXT1 dimer
4. importin
NXF1/NXT1 dimer
During the process of nuclear import, a GEF works in the:
1- cytoplasm to exchange GTP bound to Ran for GDP
2- nucleus to use GTP to release Ran from importin
3- nucleus to exchange GDP bound to Ran for GTP
4- nucleus to activate the intrinsic GTPase activity of Ran
Nucleus to exchange GDP bound to Ran for GTP
During nuclear import, a guanine exchange factor (Ran-GEF) in the nucleus facilitates the exchange of GDP for GTP on Ran, activating Ran-GTP for its role in nuclear transport.
Cytoskeleton and cell
movement
The cytoskeleton is a network of protein filaments and tubules in the cytoplasm that provides structural support, shape, and facilitates cell movement. It plays a crucial role in maintaining cell integrity, organizing cellular components, and enabling processes like migration, division, and transport
Microfilaments (Actin filaments):
Made of actin protein.
Involved in cell shape, muscle contraction, and cell movement (e.g., through amoeboid movement, filopodia, and lamellipodia).
They also contribute to cytokinesis and vesicle transport.
Intermediate Filaments
Made of various proteins (e.g., keratins, vimentin, desmin).
Provide mechanical strength to cells and tissues, helping them resist stretching and deformation.
They anchor organelles and form the nuclear lamina.
Microtubules:
Made of tubulin protein.
Form the mitotic spindle during cell division and act as tracks for intracellular transport.
Involved in the structure of cilia and flagella, which enable cell movement in some cells.
intermediate filaments
Named because they
are intermediate in size
(10 nm)
* Present throughout the
cytoplasm as well as
lining the inner nuclear
envelope
Actin – “treadmilling”
Actin can effectively move by adding subunits at one end faster than
the other
– This is an ATP-dependent process
* The rate of addition at the (+) end is much faster than at the (-) end
Muscle Structure
Muscle fiber: large, single, elongated, multinuclear cell
* Each fiber contains about 1,000 myofibrils
Myofibrils contain thick filaments
of myosin
thick filaments
* Mostly myosin
* Diameter 15 nm
* Produce the dark bands
Myofibrils contain thin filaments
of actin
Thin filaments
* Mostly actin with
some troponin and
tropomyosin
* Diameter 7 nm
* Anchored at the Z-
lines
Myosin thick filaments slide along
actin thin filaments
Muscle contraction occurs when the thin and thick filaments slide past each other
- this draws the Z disks closer together
Thick and thin filaments are interleaved so that each thick filament (myosin) is surrounded by six thin filaments (actin)
Actomyosin Cycle Summary (Muscle Contraction)
The actomyosin cycle is responsible for muscle contraction and occurs through a series of conformational changes driven by ATP binding, hydrolysis, and release. The cycle consists of four key steps:
ATP Binding: ATP binds to myosin, causing it to dissociate from actin.
ATP Hydrolysis: ATP is hydrolyzed to ADP and Pi, leading to a conformational change in myosin.
Myosin Reattachment: Myosin binds to actin at a new position, and the inorganic phosphate (Pi) is released.
Power Stroke: The release of Pi triggers the power stroke, where myosin returns to its initial conformation, pulling the actin filament along, and releasing ADP.
This cycle repeats, generating the force needed for muscle contraction.
regulation of Muscle Contraction
Troponin and Tropomyosin regulate muscle contraction by blocking myosin-binding sites on actin, preventing continuous contraction.
Nerve impulse triggers the release of Ca²⁺, which binds to troponin, causing a conformational change.
This change shifts tropomyosin, exposing the myosin-binding sites on actin, allowing muscle contraction to occur.
When comparing intermediate filaments to
microfilaments. What is INCORRECT?
A- intermediate filaments are less dynamic
B- no motor proteins “walk” on intermediate filaments
C- intermediate filaments are unpolarized
D-They use any type of energy
They use any type of energy
Actin filaments can take different physical forms
within a cell. This is possible because?
A- Actin is present in only a few cell types
B- Actin filaments grows dynamically by associating on
the +end and dissociating on the –end
C- Actin can bind GTP
D- Actin concentration is low within a cell and this
triggers the filament formation
actin filaments grow dynamically by associating on the + end and dissociating on the - end.
This dynamic growth and shrinkage of actin filaments, known as treadmilling, allows actin filaments to take various physical forms within a cell, contributing to processes like cell movement, shape changes, and division.
Myosin movements is dependent on:
A- the presence of tropomyosin
B- microtubules
C- ATP hydrolysis
D- association with troponin complex.
Myosin movement is dependent on ATP hydrolysis, which provides the energy for the conformational changes in myosin, allowing it to interact with actin and generate movement, such as in muscle contraction.
‘Enzyme’ derives from the Greek
‘enzymos’, meaning ‘leavened’.
catayst definition
A catalyst is a substance which
when present in small amounts
increases the rate of a chemical
reaction or process, but which is
chemically unchanged by the
reaction (OED)
enzyme definition
An enzyme (originally known as a
‘ferment’) is a biological catalyst.
Almost all enzymes are proteins.An enzyme is a biological catalyst that speeds up chemical reactions in living organisms without being consumed in the process. Enzymes are typically proteins and work by lowering the activation energy required for a reaction to occur, thereby increasing the reaction rate. Enzymes are highly specific to their substrates, meaning they bind to specific molecules to catalyze particular reactions.
Enzymes are central to biochemical processes.
Enzymes act in ordered
sequences of chemical reactions
known as biochemical
pathways
Enzymes have great practical importance in medicine
and industry.
Genetic diseases of humans
often involve deficiencies of
specific enzymes;
– e.g. Krabbe disease is caused by a
deficiency in galactosylceramidase,
which results in an impairment of the
growth and maintenance of myelin.
some enzymes require cofactors to function
some enzymes require
cofactors.
Cofactors can be inorganic
ions, or complex organic or
metalloorganic compounds
known as coenzymes.
A catalytically active enzyme
with its bound metal ion and/or
coenzyme is a holoenzyme.
The protein part of a
holoenzyme is the apoprotein
or apoenzyme.
The active site of an enzyme binds the substrate.
The substrate is a molecule that is
bound to the active site and acted
on by the enzyme.
The active site surface is lined with
amino acid residues with R groups
that bind the substrate.
Enzymes affect reaction rates, not equilibria.
A reaction is at equilibrium
when there is no net change in
the concentrations of reactants
or products.
An enzyme increases the rate
of a reaction but does not
affect the equilibrium
ground state
The ground state is the
contribution to the free energy of
the system by an average
molecule (S or P)
A negative DG¢°
A negative DG¢° means the
reaction is ‘favourable’ but does
not mean that S®P will occur at a
detectable rate.
The transition state is
The transition state is the point of highest energy in the reaction, where bond breakage, formation, and charge development are at a critical point. It is a fleeting state and not a stable chemical species or intermediate.
Enzymes enhance reaction rates by lowering activation energies
The rate of the reaction is
dependent on the difference
between the free energy of the
transition state and the ground
state, known as the activation
energy, DG ‡