Evaluation of the flow-dialysis technique for analysis of protein-ligand interactions: an experimental and a monte carlo study.

Abstract:

:Flow dialysis has found widespread use in determining the dissociation constant (KD) of a protein-ligand interaction or the amount of available binding sites (E0). This method has the potency to measure both these parameters in a single experiment and in this article a method to measure simultaneously the KD and E0 is presented, together with an extensive error analysis of the method. The flow-dialysis technique is experimentally simple to perform. However, a number of practical aspects of this method can have a large impact on the outcome of KD and E0. We have investigated all sources of significant systematic and random errors, using the interaction between mannitol and its transporter from Escherichia coli as a model. Monte Carlo simulations were found to be an excellent tool to assess the impact of these errors on the binding parameters and to define the experimental conditions that allow their most accurate estimation.

journal_name

Biophys J

journal_title

Biophysical journal

authors

Veldhuis G,Vos EP,Broos J,Poolman B,Scheek RM

doi

10.1016/S0006-3495(04)74259-9

subject

Has Abstract

pub_date

2004-04-01 00:00:00

pages

1959-68

issue

4

eissn

0006-3495

issn

1542-0086

pii

S0006-3495(04)74259-9

journal_volume

86

pub_type

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