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A microfluidic platform for quantitative measurements of effective protein charges and single ion binding in solution

Herling, Therese W.; Arosio, Paolo; Mueller, Thomas; Linse, Sara LU and Knowles, Tuomas P. J. (2015) In Physical Chemistry Chemical Physics 17(18). p.12161-12167
Abstract
The charge state of proteins in solution is a key biophysical parameter that modulates both long and short range macromolecular interactions. However, unlike in the case of many small molecules, the effective charges of complex biomolecules in solution cannot in general be predicted reliably from their chemical structures alone. Here we present an approach for quantifying the effective charges of solvated biomolecules from independent measurements of their electrophoretic mobilities and diffusion coefficients in free solution within a microfluidic device. We illustrate the potential of this approach by determining the effective charges of a charge-ladder family of mutants of the calcium binding protein calbindin D-9k in solution under... (More)
The charge state of proteins in solution is a key biophysical parameter that modulates both long and short range macromolecular interactions. However, unlike in the case of many small molecules, the effective charges of complex biomolecules in solution cannot in general be predicted reliably from their chemical structures alone. Here we present an approach for quantifying the effective charges of solvated biomolecules from independent measurements of their electrophoretic mobilities and diffusion coefficients in free solution within a microfluidic device. We illustrate the potential of this approach by determining the effective charges of a charge-ladder family of mutants of the calcium binding protein calbindin D-9k in solution under native conditions. Furthermore, we explore ion-binding under native conditions, and demonstrate the ability to detect the chelation of a single calcium ion through the change that ion binding imparts on the effective charge of calbindin D-9k. Our findings highlight the difference between the dry sequence charge and the effective charge of proteins in solution, and open up a route towards rapid and quantitative charge measurements in small volumes in the condensed phase. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Physical Chemistry Chemical Physics
volume
17
issue
18
pages
12161 - 12167
publisher
Royal Society of Chemistry
external identifiers
  • wos:000353767500024
  • scopus:84928910732
ISSN
1463-9084
DOI
10.1039/c5cp00746a
language
English
LU publication?
yes
id
de226a6c-2066-433f-abb9-d461fa2b0bb0 (old id 7432825)
date added to LUP
2015-06-24 15:12:59
date last changed
2017-01-01 06:09:32
@article{de226a6c-2066-433f-abb9-d461fa2b0bb0,
  abstract     = {The charge state of proteins in solution is a key biophysical parameter that modulates both long and short range macromolecular interactions. However, unlike in the case of many small molecules, the effective charges of complex biomolecules in solution cannot in general be predicted reliably from their chemical structures alone. Here we present an approach for quantifying the effective charges of solvated biomolecules from independent measurements of their electrophoretic mobilities and diffusion coefficients in free solution within a microfluidic device. We illustrate the potential of this approach by determining the effective charges of a charge-ladder family of mutants of the calcium binding protein calbindin D-9k in solution under native conditions. Furthermore, we explore ion-binding under native conditions, and demonstrate the ability to detect the chelation of a single calcium ion through the change that ion binding imparts on the effective charge of calbindin D-9k. Our findings highlight the difference between the dry sequence charge and the effective charge of proteins in solution, and open up a route towards rapid and quantitative charge measurements in small volumes in the condensed phase.},
  author       = {Herling, Therese W. and Arosio, Paolo and Mueller, Thomas and Linse, Sara and Knowles, Tuomas P. J.},
  issn         = {1463-9084},
  language     = {eng},
  number       = {18},
  pages        = {12161--12167},
  publisher    = {Royal Society of Chemistry},
  series       = {Physical Chemistry Chemical Physics},
  title        = {A microfluidic platform for quantitative measurements of effective protein charges and single ion binding in solution},
  url          = {http://dx.doi.org/10.1039/c5cp00746a},
  volume       = {17},
  year         = {2015},
}