388 Chapter 10. Enzymes and molecular machines[[Student version, January 17, 2003]]The weakly bound state M·K·Dreadily dissociates, either to M+K·DortoM·K+D.
Assumption A3 says that the free energy gain from ATP hydrolysis and phosphate release is partly
spent on a conformational change that pulls the M·K·Tcomplex out of a deep potential-energy well
to a shallower one. Similarly, A1 says that some of this free energy goes to relax the head’s grip on
its neck linker. One more geometric assumption will be explained below.Provisional model: Mechanism The proposed mechanism is summarized graphically in Fig-
ure 10.21 on page 384. This cycle is not meant to be taken literally; it just shows some of the
distinct steps in the enzymatic pathway. Initially (top-left panel of the figure), a kinesin dimer
approaches the microtubule from solution and binds one head, releasing one of its ADPs. We name
the subsequent states in the ATP hydrolysis cycle by abbreviations describing the state of the head
labeled “Ka.”
E:The top-middle panel shows the dimer with Kabound to the microtubule. The other, free
head Kbcannot reach any binding site, because its tether is too short; the sites are separated
byafixed distance along the rigid microtubule.ES 1 ,ES′ 1 :Kabinds an ATP molecule from solution. Its neck linker then docks onto its head, biasing
the other head’s random motion in the forward direction (assumption A2). Schnitzer and
coauthors assumed that Kb’s interactions with the microtubule effectively give it a weak
energy landscape, so that it hops between two distinct states ES 1 and ES′ 1.
ES 2 :The chains joining the two heads will have entropic elasticity, as discussed in Chapter 9.
Being thrown forward by Ka’s neck linker greatly increases the probability that Kb’s tether
will momentarily stretch far enough to reach the next binding site. It may bind weakly, then
detach, many times.
ES 3 :Eventually, instead of detaching, Kbreleases its ADP and binds strongly to the microtubule.
Its stretched tether now places the dimer under sustained strain. Both heads are now tightly
bound to the microtubule, however, so the strain does not pull either one off.
EP:Meanwhile, Kasplits its ATP and releases the resulting phosphate. This weakens its binding
to the microtubule (assumption A3). The strain induced by the binding of Kbthen biases Ka
to unbind from the microtubule (rather than releasing its ADP).
E:The cycle is now ready to repeat, with the roles of Kaand Kbreversed. The kinesin dimer
has made one 8nmstep and hydrolyzed one ATP.
The assumptions made above ensure thatfreekinesin (not bound to any microtubule) does not
waste any of the available ATP, as observed experimentally. According to assumption A3, free
kinesin will bind and hydrolyze ATP at each of its two heads, then stop, because the resulting
ADPs are both tightly bound in the absence of a microtubule.
Our model is certainly more complicated than the S-ratchet device imagined in Section 10.2.2!
But we can see how our assumptions implement the necessary conditions for a molecular motor
found there (Idea 10.9a,b on page 369):
- The forward flip induced by neck linker binding (assumption A1), together with the
polarity of the microtubule, creates the needed spatial asymmetry. - The tight linkage to the hydrolysis of ATP creates the needed out-of-equilibrium con-
dition, since the cell maintains the reaction quotientcATP/(cADPcPi)atalevel much
higher than its equilibrium value.