Conformational changes of calmodulin upon Ca2+ binding studied with a microfluidic mixer
Abstract
A microfluidic mixer is applied to study the kinetics of calmodulin conformational changes upon Ca2+ binding. The device facilitates rapid, uniform mixing by decoupling hydrodynamic focusing from diffusive mixing and accesses time scales of tens of microseconds. The mixer is used in conjunction with multiphoton microscopy to examine the fast Ca2+-induced transitions of acrylodan-labeled calmodulin. We find that the kinetic rates of the conformational changes in two homologous globular domains differ by more than an order of magnitude. The characteristic time constants are ≈490 μs for the transitions in the C-terminal domain and ≈20 ms for those in the N-terminal domain of the protein. We discuss possible mechanisms for the two distinct events and the biological role of the stable intermediate, half-saturated calmodulin.
Additional Information
© 2008 by The National Academy of Sciences of the USA. Contributed by Watt W. Webb, November 14, 2007 (sent for review October 12, 2007). We thank Mark A. Williams for editorial assistance. This work was supported by the Cornell Nanobiotechnology Center, a Science and Technology Center Program of the National Science Foundation, under Agreement ECS-9876771; National Institute of Biomedical Imaging and Bioengineering/National Institutes of Health Grant 9 P41 EB001976; and National Institute of General Medical Sciences Grant P01-GM066275. All fabrication work was done at the Cornell NanoScale Science and Technology Facility, which is supported by the National Science Foundation, Cornell University, and industrial affiliates.Attached Files
Published - PARpnas08.pdf
Supplemental Material - 10810SuppText.pdf
Supplemental Material - PARpnas08figs.pdf
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Additional details
- PMCID
- PMC2206572
- Eprint ID
- 12191
- Resolver ID
- CaltechAUTHORS:PARpnas08
- ECS-9876771
- NSF
- 9 P41 EB001976
- NIH
- P01-GM066275
- NIH
- Cornell University
- National Institute of Biomedical Imaging and Bioengineering
- National Institute of General Medical Sciences
- Created
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2008-11-13Created from EPrint's datestamp field
- Updated
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2021-11-08Created from EPrint's last_modified field