Chemistry Reference
In-Depth Information
with an average of 2.5 steps at low loads (0
-
0.5 pN) and from one to four with an
average of 1.6 steps at high loads (0.5
-
2 pN).
2.3.4
Nature of Sub-steps
The steppingmotion of amyosin head is not always smooth and sometimesmoves in
the opposite direction along an actin
filament. The size of the steps was 5.5 nm for
steps in both the forward and backward directions. This step size coincided with the
distance between adjacent actin monomers in one strand of an actin
filament [21].
Furthermore, the number of steps ranged randomly from one to
five during nomore
than one ATP hydrolysis cycle, i.e. the 5.5-nm steps were not tightly coupled to the
ATP hydrolysis cycle. The stochastic features of this steppingmotion and the step size
strongly suggest that themyosin headwalks or slides along the actinmonomer repeat
using Brownianmotion. Because themajority of steps occurred in one direction, they
should not result from pure thermal diffusion but rather be biased in one direction
(forward).
In order to clarify this point quantitatively, the number of forward (Nf)
f
) and
backward (N
b
) steps were counted at low force levels and the (N
f
/N
b
) ratio was
calculated. The ratio of the number of forward and backward steps near zero force
(0
-
0.5 pN) was 55/9
6 (Figure 2.5d). Thus the probability that the 55 forward steps
out of total 64 steps were due to pure Brownian motion could be calculated as
55
9
64
!
55
!
9
!
1
2
1
2
10
9
:
In this calculation, it was assumed that no external force was applied to the myosin.
However, because the myosin underwent steps against a force, the actual probability
should be even less than the above result. Therefore, the observed steps cannot be due
to pure Brownian motion. Rather, myosin Brownian steps are likely biased towards
the forward direction.
2.3.5
Comparing the Actions of Individual Myosin Motors with those of Muscle
To quantitatively compare the mechanical properties of individual actomyosin
motors with those of muscle, their force
-
velocity curve was investigated. Displace-
ments started at various force levels due to thermal
fluctuations of the needle causing
steps to also take place at various force levels. Figure 2.6 shows the force
-
velocity
curve obtained from the step size and the dwell time of single myosin heads. The
velocity did not reach zero but a positive value at a large positive force, which is
different from that of muscle. This ismost likely because the backward steps were not
considered in the analysis. Backward steps took place more frequently at larger
forces, thus the backward steps need to be considered when calculating the velocity of
individual myosin heads. The anisotropy of the stepping direction was de
ned as
(N
f
N
b
)/(N
f
รพ
N
b
), where N
f
and N
b
are the number of steps in the forward and