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Electrostatic contributions to the kinetics and thermodynamics of protein assembly

Dell'Orco, D; Xue, Wei-Feng LU ; Thulin, Eva LU and Linse, Sara LU (2005) In Biophysical Journal 88(3). p.1991-2002
Abstract
The role of electrostatic interactions in the assembly of a native protein structure was studied using fragment complementation. Contributions of salt, pH, or surface charges to the kinetics and equilibrium of calbindin D-9k reconstitution was measured in the presence of Ca2+ using surface plasmon resonance and isothermal titration calorimetry. Whereas surface charge substitutions primarily affect the dissociation rate constant, the association rates are correlated with subdomain net charge in a way expected for Coulomb interactions. The affinity is reduced in all mutants, with the largest effect (260-fold) observed for the double mutant K25E+K29E. At low net charge, detailed charge distribution is important, and charges remote from the... (More)
The role of electrostatic interactions in the assembly of a native protein structure was studied using fragment complementation. Contributions of salt, pH, or surface charges to the kinetics and equilibrium of calbindin D-9k reconstitution was measured in the presence of Ca2+ using surface plasmon resonance and isothermal titration calorimetry. Whereas surface charge substitutions primarily affect the dissociation rate constant, the association rates are correlated with subdomain net charge in a way expected for Coulomb interactions. The affinity is reduced in all mutants, with the largest effect (260-fold) observed for the double mutant K25E+K29E. At low net charge, detailed charge distribution is important, and charges remote from the partner EF-hand have less influence than close ones. The effects of salt and pH on the reconstitution are smaller than mutational effects. The interaction between the wild-type EF-hands occurs with high affinity (K-A = 1.3 x 10(10) M-1; K-D = 80 pM). The enthalpy of association is overall favorable and there appears to be a very large favorable entropic contribution from the desolvation of hydrophobic surfaces that become buried in the complex. Electrostatic interactions contribute significantly to the affinity between the subdomains, but other factors, such as hydrophobic interactions, dominate. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biophysical Journal
volume
88
issue
3
pages
1991 - 2002
publisher
Cell Press
external identifiers
  • wos:000227494600046
  • pmid:15596501
  • scopus:21244433239
ISSN
1542-0086
DOI
10.1529/biophysj.104.049189
language
English
LU publication?
yes
id
0f09ce5a-aeb7-4af6-8c58-566ec3aaec4a (old id 151555)
date added to LUP
2007-06-29 09:07:47
date last changed
2017-07-23 03:55:51
@article{0f09ce5a-aeb7-4af6-8c58-566ec3aaec4a,
  abstract     = {The role of electrostatic interactions in the assembly of a native protein structure was studied using fragment complementation. Contributions of salt, pH, or surface charges to the kinetics and equilibrium of calbindin D-9k reconstitution was measured in the presence of Ca2+ using surface plasmon resonance and isothermal titration calorimetry. Whereas surface charge substitutions primarily affect the dissociation rate constant, the association rates are correlated with subdomain net charge in a way expected for Coulomb interactions. The affinity is reduced in all mutants, with the largest effect (260-fold) observed for the double mutant K25E+K29E. At low net charge, detailed charge distribution is important, and charges remote from the partner EF-hand have less influence than close ones. The effects of salt and pH on the reconstitution are smaller than mutational effects. The interaction between the wild-type EF-hands occurs with high affinity (K-A = 1.3 x 10(10) M-1; K-D = 80 pM). The enthalpy of association is overall favorable and there appears to be a very large favorable entropic contribution from the desolvation of hydrophobic surfaces that become buried in the complex. Electrostatic interactions contribute significantly to the affinity between the subdomains, but other factors, such as hydrophobic interactions, dominate.},
  author       = {Dell'Orco, D and Xue, Wei-Feng and Thulin, Eva and Linse, Sara},
  issn         = {1542-0086},
  language     = {eng},
  number       = {3},
  pages        = {1991--2002},
  publisher    = {Cell Press},
  series       = {Biophysical Journal},
  title        = {Electrostatic contributions to the kinetics and thermodynamics of protein assembly},
  url          = {http://dx.doi.org/10.1529/biophysj.104.049189},
  volume       = {88},
  year         = {2005},
}