Determination of dissociation constants between polyelectrolytes and proteins by affinity capillary electrophoresis

Maria Anderot, Mikael Nilsson, Ákos Végvári, Eva Horn Moeller, Marco van de Weert, Roland Isaksson

Research output: Contribution to journalArticlepeer-review

22 Citations (SciVal)

Abstract

In this manuscript we report the binding affinity between two model proteins, human serum albumin (HSA) and ribonuclease A (RNase A), and negatively charged polyelectrolytes, two different heparin fractions and dextran sulfate, by means of partial filling and affinity capillary electrophoresis. The apparent dissociation constants, Kd, obtained by use of the partial-filling method, between HSA and heparin (17 kDa), heparin (3 kDa) and dextran sulfate (8 kDa) were 33 and 307 μM, respectively. A new method was developed to determine affinities that take in account different migration directions between the protein and the polyelectrolyte, which was required to study RNase A. By use of this affinity capillary electrophoresis two Kd values were observed for the interaction between RNase A and heparin 17 kDa, yielding a high affinity binding with Kd1 0.0075 μM, and a lower affinity binding with Kd2 8.7 μM. For dextran sulfate 8 kDa these Kd values were 0.027 and 10.4 μM, respectively. Heparin 3 kDa only showed a single Kd value of 0.52 μM. The results show that the magnitude of the binding affinity depends on the type of polyelectrolyte and its molecular weight.
Original languageEnglish
Pages (from-to)892-896
Number of pages5
JournalJournal of Chromatography. B
Volume877
Issue number10
DOIs
Publication statusPublished - 2009 Apr 1

Subject classification (UKÄ)

  • Medical Engineering

Keywords

  • dissociation constant
  • heparin
  • dextran sulfate
  • human serum albumin
  • capillary electrophoresis
  • ribonuclease A
  • RNase
  • affinity binding
  • partial filling
  • polyelectrolytes

Fingerprint

Dive into the research topics of 'Determination of dissociation constants between polyelectrolytes and proteins by affinity capillary electrophoresis'. Together they form a unique fingerprint.

Cite this