Journal
BIOPHYSICAL JOURNAL
Volume 121, Issue 9, Pages 1715-1726Publisher
CELL PRESS
DOI: 10.1016/j.bpj.2022.03.029
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Funding
- Clemson University's R-Initiative CU-MRI grant
- Department of Physics and Astronomy, the College of Science, the Creative Inquiry Program
- Clemson University Professional Internship and Co-op Program (UPIC)
- National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health [R15AI137979]
- National Institute of General Medical Sciences (NIGMS) of the National Institutes of Health [P20GM109094]
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The dynein family of motor proteins plays diverse roles in eukaryotic cells and their processivity is regulated by both specific salt bridges and long-range electrostatic interactions.
The dynein family of microtubule minus-end-directed motor proteins drives diverse functions in eukaryotic cells, including cell division, intracellular transport, and flagellar beating. Motor protein processivity, which characterizes how far a motor walks before detaching from its filament, depends on the interaction between its microtubule-binding domain (MTBD) and the microtubule. Dynein's MTBD switches between high- and low-binding affinity states as it steps. Significant structural and functional data show that specific salt bridges within the MTBD and between the MTBD and the microtubule govern these affinity state shifts. However, recent computational work suggests that nonspecific, long-range electrostatic interactions between the MTBD and the microtubule may also play an important role in the processivity of dynein. To investigate this hypothesis, we mutated negatively charged amino acids remote from the dynein MTBD-microtubule-binding interface to neutral residues and measured the binding affinity using microscale thermophoresis and optical tweezers. We found a significant increase in the binding affinity of the mutated MTBDs for microtubules. Furthermore, we found that charge screening by free ions in solution differentially affected the binding and unbinding rates of MTBDs to microtubules. Together, these results demonstrate a significant role for long-range electrostatic interactions in regulating dynein-microtubule affinity. Moreover, these results provide insight into the principles that potentially underlie the biophysical differences between molecular motors with various processivities and protein-protein interactions more generally.
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