Cytoskelton: Myosin Flashcards
Actin works with motor proteins: myosins
Have a globular motor domain which uses ATP to move (conformational change of the motor domain)
Many function as dimers
All myosins EXCEPT myosin VI move toward the plus end of actin
Actin + myosin
Myosin 2
- conventional myosin
- globular (rounded motor domains at n-terminus)
- Long coiled-coiled domain where myosin proteins interact with each other
- 2 light chains per heavy chain (4 per myosin)
Myosin 5: moves organelles along actin
- has a very long step size
- carries vesicles, Er, mRNA, organelles like mitochondria
In vitro assay to study assay
- myosin molecules are bound top a glass slides
- fluorescently labeled actin filaments will be added to the slides and bind to myosin
- can measure speed and direction of movement
Steps of mysosin movement
- Rigor confirmation: Myosin head attached to actin filament (without ATP) so very stiff -> rigor mortis
- Release: ATP binds to the head domain
- Clocked: cleft around ATP binding site closes and leads to movement in the lever arm -> displaces the motor head -> ATP hydrolyzed to ADP -> ADP has affinity to bind to actin
- Force-generating: head domain binds to actin weakly, which causes phosphate group to be released -> which causes tight binding and triggers the power stroke ->
Attached: go back to rigor confirmation
Actomyosin contractility in skeletal muscle: muscle cells
- muscle cells are multinucleate, and have bundles called myofibrils within the that are mostly made of actin and myosin
- can use TEM to view this
- organization of carsocmeres in myofibrils is repeated structural units
Dark bands in the middle: myosin (m line)
Light band (z-disc): actin
Actomyosin contractility in skeletal muscle: myosin bundles
- Myosin 2 dimers can self organize into bipolar thick filaments
- heads of myosin on each end face the same in opposite ways
- interact via coiled-coiled domain
- bare zone is m-line
Actin and assoaciacted proteins
- Tropomodulin: binds to actin’s minus end and cap it
- nebulin: to bind along filament, controls actin length
- Cap z on plus size
ACTIN IS NOT DYNAMIC IN THIS CASE, because of the caps, so it will stay the size
Titin (protein)
- anchors myosin to the z-disc
Contractility
Filaments is repeating patters
- shortening of sarcomere WO shortening actin or myosin
How is contraction regulated?
- AP from neuron that will excite it
- Excitation is spread quickly along plasma membrane of the cell and through t-tubules (that link plasma membrane with regions throughout the muscle)
- Signal will be relayed to sarcoplasmic reticulum (modified ER) that surrounds myofibrils
- Voltages gated calcium channel on polarized t-tubule membrane, excitation will cause Ca2+ influx into the cytosol
- Also causes ca2+ release from sarcoplasmic reticulum to the the cytoplasm
- Quick rise in calcium, high Ca2+ will be reversed through calcium pumps
Impact of myosin: modifies actin-binding proteins : changes availability of actin to myosin
Tropomyosin: accessory protein of actin in skeletal muscle
Troponin: Tropomyosin-binding, inhibitory, and Ca2+ binding
At rest: T and I pull tropomyosin in position to block myosin interaction with actin
WITH HIGH CA2+ -> C which binds to 4 Ca2+ causes troponin I to release actin -> tropomyosin slides into another position, allowing myosin to access actin filaments
Smooth muscle: how contractility work
- myosin always in skeletal muscle but not in smooth muscle
Smooth muscles are a part of stomachs, uterus, blood vessels
1. Sarcoplasmic reticulum is still going to release CA2+ after receiving a signal from a neuron
2. Calcium will bind to calmodulin instead troponin C, to change conformation of protein
3. Calmodulin will activate MLCK (myosin light chain kinase) which will phosphorylate myosin light chain (important for regulation)
4. Light chains, once phosphorylated will change conformation of myosin dimer, once released it can form bipolar filaments
Cell movement depends on actin polymerization
Top: actin filaments in red (shown by phalloidin) and arp2/3 complex in green
Branched actin filaments in the lamellipodia are essential for rapid cellular movement
Cell motility
- arp2/3 builds branched actin that pushes membrane
- actin network undergoes treadmiling
- depolymerization at sites well behind the leading edge (cofilin)
- actin capping at a steady rate
