1. Motor Proteins - Introduction Part 1
Biochemistry 4000
Dr. Ute Kothe

2. Motor Proteins Motor Proteins convert chemical energy into motion.
chemical energy is derived from ATP hydrolysis
motion is generated by conformational changes depending on the
bound nucleotide
Myosin Kinesin Dynein

3. Motor Protein Function
Myosin (18 known classes, 40 different myosins in humans)
Movement along actin fibres
Muscle movement
Cytokinesis (cytoplasmic division, tightening of contractile ring)
Transport of cargo along microfilaments (vescicles etc.)
Kinesin (16 classes)
Movement along microtubule tracks, usually to (+) end
Transport of cargos: vesicles, organelles, cytosolic components such as mRNAs & proteins, chromosomes
Dynein (12 mammalian dyneins)
Movement along microtubule tracks, to (-) end, i.e. cell center
Cytoplasmic dyneins: transport of cargos such as vesicles
Axonemal dyneins: Movement of cilia and flagella

4. Tubulin building block of microtubules
heterodimer of closely related Tubulin a & b
G proteins: N-terminal residues fold into G domain-like structure
a-tubulin’s GTP buried at subunit interface, nonexchangable, not hydrolyzed
b-tubulin’s GTP is solvent exposed until tubulin dimers polymerize
upon polymerization, a-tubulin from adjacent dimer provides catalytic residue to hydrolyze b-tubulin’s GTP; resulting GDP is nonexchangeable unless tubulin dissociates from microtubule
Tubulin b
+ GDP
Tubulin a
+ GTP
Voet Fig

5. Microtubules
Tubulins interact head to tail to form a long protofilament
Protofilaments align side by side in curved sheet
Sheet of 13 (9-16) protomers closes on itself to form microtubule
Microtubule lengthens by addition of tubulins to both ends (preferentially to + end, i.e. the end terminating in b-tubulins)
Voet Fig

6. Structure of the Axoneme
Bundle of microtubules called axoneme
coated by plasma membrane
Forms eukaroytic flagella & cilia
Voet Fig

7. Dynein 1 or more heavy chains (motor domain)
several intermediate and light chains
motor domain is 7-membered ring, ATP-hydrolyzing unit
coiled-coil extension forms stalk that interacts via globular domain with microtubules
long stem (with intermediate and light chains) binds cargo
Valle, Cell 2003; Voet Fig

8. Conventional Kinesin
two identical heavy chains forming two large globular heads attaching to microtubules and a coiled-coil
two identical light chains interacting wit cargo
transports vesicles and organelles in (-) to (+) direction (towards cell periphery)
Voet Fig

9. Kinesin Structure
Globular head: tubulin-binding site & nucleotide binding site
flexible neck linker
a-helical stalk leading into coiled-coil
ATP hydrolysis triggers conformational change in neck linker via 2 switch regions:
When ATP is bound, neck linker docks with catalytic core
Upon ATP hydrolysis, the neck linker “unzips”
Voet Fig & 96 & 97

10. Kinesin Cycle
Voet Fig

11. Hand-over-Hand Mechanism
ATP-bound state: strong microtuble binding
ADP-bound state: weak microtubule binding
ATP binds to leading head - globular kinesin head which is already bound to the microtubule and oriented towards (+) end
Neck linker of leading head “zips up” agains catalytic core
trailing head is thrown forward (trailing head has bound ADP and reduced affinity to microtubule):
Trailing head swings by ~ 160 Å, net movement of dimeric kinesin is ~ 80 Å = length of one microtubule dimer
ATP in new trailing head is hydrolyzed & phosphate released: affinity for microtuble decreases
ADP in new leading head dissociates
Two heads work in a coordinated fashion
ATP binding to leading head induces power stroke

12. Processivity
Kinesin is highly processive: it takes several 100 steps on a microtubule without detaching or sliding backwards!
How?
coordinated, but out of phase ATP cycle in both heads
one head is always firmly attached to microtubule
Movie demonstrating kinesins processivity:

13. Dynein 1 or more heavy chains (motor domain)
several intermediate and light chains
motor domain is 7-membered ring, ATP-hydrolyzing unit
coiled-coil extension forms stalk that interacts via globular domain with microtubules
long stem (with intermediate and light chains) binds cargo
Valle, Cell 2003; Voet Fig