Monte Carlo analysis of neck linker extension in kinesin molecular motors - PubMed (original) (raw)
Monte Carlo analysis of neck linker extension in kinesin molecular motors
Matthew L Kutys et al. PLoS Comput Biol. 2010.
Abstract
Kinesin stepping is thought to involve both concerted conformational changes and diffusive movement, but the relative roles played by these two processes are not clear. The neck linker docking model is widely accepted in the field, but the remainder of the step--diffusion of the tethered head to the next binding site--is often assumed to occur rapidly with little mechanical resistance. Here, we investigate the effect of tethering by the neck linker on the diffusive movement of the kinesin head, and focus on the predicted behavior of motors with naturally or artificially extended neck linker domains. The kinesin chemomechanical cycle was modeled using a discrete-state Markov chain to describe chemical transitions. Brownian dynamics were used to model the tethered diffusion of the free head, incorporating resistive forces from the neck linker and a position-dependent microtubule binding rate. The Brownian dynamics and chemomechanical cycle were coupled to model processive runs consisting of many 8 nm steps. Three mechanical models of the neck linker were investigated: Constant Stiffness (a simple spring), Increasing Stiffness (analogous to a Worm-Like Chain), and Reflecting (negligible stiffness up to a limiting contour length). Motor velocities and run lengths from simulated paths were compared to experimental results from Kinesin-1 and a mutant containing an extended neck linker domain. When tethered by an increasingly stiff spring, the head is predicted to spend an unrealistically short amount of time within the binding zone, and extending the neck is predicted to increase both the velocity and processivity, contrary to experiments. These results suggest that the Worm-Like Chain is not an adequate model for the flexible neck linker domain. The model can be reconciled with experimental data if the neck linker is either much more compliant or much stiffer than generally assumed, or if weak kinesin-microtubule interactions stabilize the diffusing head near its binding site.
Conflict of interest statement
The authors have declared that no competing interests exist.
Figures
Figure 1. Kinesin Chemomechanical Pathway.
Working model for the Kinesin-1 chemomechanical pathway based on previous experimental work. Nucleotide abbreviations are as follows: T = ATP, D = ADP, DP = ADP.Pi, φ = No nucleotide. For clarity, ADP bound to tethered head in states 2–4 is not shown. In State 2 the tethered head diffuses, tethered by both neck linker domain, while in states 3 and 4 the neck linker domain of the bound head is docked, leading to a displacement of the tethered head towards the next binding site. State 5 represents motor detachment. Note that the number of steps per interaction (motor processivity) can be approximated by kattach/kunbind.
Figure 2. Kinesin Structural Models.
A: Comparison of neck linker structures before and after docking. In state preceding ATP binding (left) the tether between the heads consists of both neck linkers (28 amino acids) with no forward bias (initial position 0 nm). Upon nucleotide binding (right), the rear neck linker docks to its motor domain, providing a 4.1 nm bias toward the microtubule plus-end. At this point the free motor head is tethered only by its 14 amino acid neck linker. The microtubule binding zone (7.2–9.2 nm, grey box) is defined as a region within 1 nm of the binding site. The motor is also permitted to bind to a site 8.2 nm to the rear (not shown), but this rarely occurs. B: Kinesin-1 force-extension profile from molecular dynamics simulations. Solid line shows fit to WLC with Lp = 0.7 nm and dashed line shows fit to WLC with Lp = 2 nm; both use Lc of 0.364 nm per amino acid as described in text. Molecular dynamics results adapted from Hariharan and Hancock . C: Force extension profiles of the neck linker domain shown for the Increasing Stiffness Model (dashed line), Constant Stiffness Model (dotted line) and Reflecting Model (solid line). Arrows represent the reflecting barrier characteristic of the Reflecting Model.
Figure 3. Stationary Distribution Profile of Tethered Head.
Stationary positional distribution of the tethered Kinesin-1 motor domain during its diffusive search using the Increasing Stiffness neck linker model. Dotted line shows state before ATP binding (State 2 in Figure 1) where both neck linkers are disordered and there is no positional bias of the tethered head. Solid line shows state following ATP binding (States 3 and 4 in Figure 1) where docking of one neck linker causes a 4 nm displacement toward the microtubule plus-end and diffusion is tethered by remaining neck linker. Note that before neck linker docking the free head cannot reach the next microtubule binding site (grey zone), while after neck linker docking the free head spends only a small fraction of the time (<1%) near the binding site.
Figure 4. Stationary Distributions for All Models.
Stationary distributions for the Increasing Stiffness, Constant Stiffness, and Reflecting models of the neck linker domain. The position of the free head of Kinesin-1 and Kinesin-1+DAL was simulated by setting kattach to zero. A 4 nm bias resulting from ATP binding and neck linker docking is assumed in all cases.
Figure 5. Possible Resolutions to the Increasing Stiffness Model.
If the neck linker is considerably stiffer than estimated from WLC models (perhaps stabilized through interactions with the core motor domain), then it would act more like a pivoting rod. Thus, the tethered head would diffuse in the vicinity of its binding site. Extending the neck linker would be expected to position the tethered head beyond its binding site, slowing the rate of attachment. Alternatively, the tethered head may be stabilized near its microtubule binding site by weak electrostatic interactions with the microtubule that counteract the restoring force of the neck linker tether. ADP release would then trigger strong binding to the microtubule.
References
- Hirokawa N, Takemura R. Kinesin superfamily proteins and their various functions and dynamics. Exp Cell Res. 2004;301:50–59. -PubMed
- Hancock WO, Howard J. Kinesin: Processivity and chemomechanical coupling. In: Schliwa M, editor. Molecular Motors. Weinheim, Germany: Wiley-VCH; 2003. pp. 243–269.
- Rice S, Lin AW, Safer D, Hart CL, Naber N, et al. A structural change in the kinesin motor protein that drives motility. Nature. 1999;402:778–784. -PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources