16-8 The movements of single motor-protein molecules can be analyzed directly. Using polarized laser light, it is pos- sible to create interference patterns that exert a centrally directed force, ranging from zero at the center to a few piconewtons at the periphery (about 200 nm from the cen- ter). Individual molecules that enter the interference pat- tern are rapidly pushed to the center, allowing them to be captured and moved at the experimenter's discretion. Using such "optical tweezers," single kinesin molecules can be positioned on a microtubule that is fixed to a cover- slip. Although a single kinesin molecule cannot be seen optically, it can be tagged with a silica bead and tracked indi- rectly by following the bead (Figure Q16-2A). In the absence of ATP, the kinesin molecule remains at the center of the interference pattern, but with ATP it moves toward the plus end of the microtubule. As kinesin moves along the micro- tubule, it encounters the force of the interference pattern, which simulates the load kinesin carries during its actual function in the cell. Moreover, the pressure against the silica bead counters the effects of Brownian (thermal) motion, so that the position of the bead more accurately reflects the position of the kinesin molecule on the microtubule. Traces of the movements of a kinesin molecule along a microtubule are shown in Figure Q16-2B. 16- ma 16- sia vic mu sli pr sic m m fil tension (% of maximum) (A) EXPERIMENTAL SETUP silica bead microtubule distance (nm) (B) POSITION OF KINESIN 138 80- trace 1 60- 40- to s kinesin 20. 0- 0 2 6 time (seconds) 8 Figure Q16-2 Movement of kinesin along a microtubule (Problem 16-8). (A) Experimental setup with kinesin linked to a silica bead, moving along a microtubule. (B) Position of kinesin (as visualized by position of silica bead) relative to center of interference pattern, as a function of time of movement along the microtubule. The jagged nature of the trace results from Brownian motion of the bead. A. As shown in Figure Q16-2B, all movement of kinesin is in one direction (toward the plus end of the microtubule). What supplies the free energy needed to ensure a unidirec- tional movement along the microtubule? B. What is the average rate of movement of kinesin along the microtubule? C. What is the length of each step that a kinesin takes as it moves along a microtubule? D. From other studies it is known that kinesin has two globular domains that each can bind to ẞ-tubulin, and that kinesin moves along a single protofilament in a micro- tubule. In each protofilament the ẞ-tubulin subunit repeats at 8-nm intervals. Given the step length and the interval between ẞ-tubulin subunits, how do you suppose a kinesin molecule moves along a microtubule? E. Is there anything in the data in Figure Q16-2B that tells you how many ATP molecules are hydrolyzed per step?