Download The cytoskeletal system, motor proteins Cyto + SKELETON

29/11/2011
The cytoskeletal system, motor
proteins
20.11.2011.
Cyto + SKELETON
• The cellular "scaffold" or "skeleton" within the cytoplasm
of the eukaryotes (prokaryotic cytoskeleton!? (1991)).
• It is composed of proteins.
• Functions:
− maintains cell shape (stability)
− protect the cell
− enables cellular motion (using structures such as
flagella, cilia and lamellipodia)
− intracellular transport (the movement of vesicles and
organelles)
− endocytosis and exocytosis
− cell division
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The cytoskeleton is made up of 3 components.
microtubules
cytoskeleton
intermediate
filaments
microfilaments
http://www.pdn.cam.ac.uk/staff/roper/kr_images.html
http://www.microscopyu.com/articles/fluorescence/filtercubes/yfp/yfphyq/stains/yfpcy2keratinptk2cells.html
http://www.lajollaneuroscience.org/sr/homepage/cell/scientific_art_gallery/pasquale.htm
Cytoskeletal filament arrangements
• filament
• bundles (parallel array of filaments – e.g. microvillus)
• network (loosley packed, crisscross filaments)
– 2D planar (close to plasma and nuclear
membranes)
– 3D (within the cell)
Cross-linking proteins !!! (connect and stabilize)
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Microfilaments / Actin
~ 7-10 nm in diameter
most concentrated just beneath the cell membrane
Functions:
• force production (contractile and protrusive forces)
• cell movement (20µm/sec.)
• wound healing
• defend against infection
• maintaining cellular shape
• participation in some cell-to-cell or cell-to-matrix
junctions (signal transduction)
• important role in cytokinesis
• muscular contraction (myosin)
Microfilaments - actin
F-actin
(polymer – filamentous)
ATP
37nm
adenosine triphosphate
G-actin (monomer - globular)
G-aktin (42.3kDa)
~ 375 amino-acid
6.7 x 4 x 3.7 nm
7-10nm
Double stranded, right handed helix
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Actin – nucleotide hidrolysis
-
+
-
+
D D D D D D D T
D
T
D D D D D D D T
T
+
The ratio of the actin monomers inside the actin filaments (%)
Actin polymerisation
nucleation
(„lag” phase)
elongation
(increasing the size)
„steady-state”
(dynamic equlibrium)
100
Critical concentration (non-polymerising actin fraction)
+
-
Formation of oligomers
0
(RATE LIMITING STEP)
time
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„Tread-milling”
shrinking end
-
+
growing end
-
+
+
-
-
+
-
+
Movement of the filament
Intermediate filaments
8 to 12 nanometers in diameter
Ropelike filaments
more stable than actin filaments (easy to bend hard to break)
Functions:
•maintenance of cell-shape by bearing tension.
•organize (STABILISE) the internal three-dimensional structure of
the cell (ceratin).
•supporting and anchoring the position of the organelles in the
cytosol (vimentin)
•structural components of the nuclear lamina (lamins) and
sarcomeres (desmin)
•They involved in some cell-cell and cell-matrix junctions.
Types:
• vimentins (the common structural support of many cells)
• keratin, found in skin cells, hair and nails
• neurofilaments of neural cells (NF-L, NF-M)
• Lamin (structural support to the nuclear envelope)
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Intermediate filaments
Polymerisation:
• Monomer
• Dimer
• Tetramer
• two tetramers….filamentum
No polarity (no + or - end)!
Microtubules
Hollow cylinders (tubes) of about 25 nm in diameter (lumen =
approximately 15nm in diameter)
Contains 13 protofilaments (polymers of alpha and beta tubulin).
They have a very dynamic behaviour, binding GTP for
polymerization.
They are commonly organized by the centrosome
(MTOC:Microtubule Organizing Center).
Functions:
• intracellular transport (associated with dyneins and kinesins).
• the mitotic spindle (cell division).
• connection with IC organelles (ER, mitochondrion)
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Microtubules - tubulin
β-tubulin
GTP v. GDP
25nm
dimer
+
GTP
„cap”
GDP
protofilament
GTP
α-tubulin
-
growth
shrinkage
GTP: Guanosine-triphosphate
Actin-binding proteins
Functions:
• Sequestering monomers (profilin – thymosin)
actin cc. in the non-muscle cells ~50-200µM ~ 50% monomer
• facilitate polymerisation (Arp2/3; formin)
• inhibit polymerisation („capping protein”)
• cutting (fragmentation, „digestion”) the polymers (gelsolin)
• Crosslinkig (spectrin, fimbrin, α-actinin)
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Thymosin - profilin
Actin-thymosin complex
works against the filament formation
-
+
free monomers
-
+
Actin-profilin complex
facilitate the filament formation
ARP2/3
70o
Stick to the side of a filament
Start the polymerisation (growing) of new filaments
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Microtubule binding proteins
• Sequestering („stathmin”)
• Stabilizing („MAP:Microtubule Associated Proteins”)
Intermedier filament binding proteins
• crosslinking (plectin)
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Motor proteins
Motor proteins
• They can bind to specific filaments
• They hydrolyze ATP (chemical energy)
• They can move along filaments (kinetic
energy)
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Types of motor proteins
1. Actin-based: myosins
Conventional (miozin II – 1864: Wilhelm Kühne) and nonconventional myosins
Myosin families: myosin I-XVIII
2. Microtubule based motors
a. Dynein (1965: Gibbons and Rowe)
Flagellar and cytoplasmic dyneins. Mw~500kDa
They move towards the minus end of MT
b. Kinesin (1985: Ron Vale)
Cytoskeletal kinesins
Neurons, cargo transport along the axons
Kinesin family: conventional kinesins + isoforms. Mw~110 kDa
They move towards the minus end of MT
3. Nucleic acid based
DNA and RNA polymerases
They move along a DNA and produce force
Motor proteins
• “Walk” or slide along cytoskeletal fibers
– Myosin on microfilaments
– Kinesin and dynein on microtubules
• Use energy from ATP hydrolysis
• Cytoskeletal fibers:
– Serve as tracks to carry organelles or vesicles
– Slide past each other
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Common properties
N
1. Structure
C
N-terminal globular head:
motor domain, nucleotide binding and hydrolysis
specific binding sites for the corresponding filaments
C-terminal: structural and functional role (e.g. myosins)
2. Mechanical properties, function
In principle: cyclic function and work
Motor -> binding to a filament -> force -> dissociation -> relaxation
1 cycle requires 1 ATP hydrolysis
They can either move or produce force
The end!
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