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Permanent link (DOI): https://doi.org/10.7939/R3930P741

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Dissociation kinetics of the streptavidin-biotin interaction measured using direct electrospray ionization mass spectrometry analysis Open Access

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Author or creator
Deng, Lu
Kitova, Elena N.
Klassen, John S.
Additional contributors
Subject/Keyword
Protein–ligand complexes
Electrospray ionization mass spectrometry
Rate constants
Type of item
Journal Article (Published)
Language
English
Place
Time
Description
Dissociation rate constants (koff) for the model high affinity interaction between biotin (B) and the homotetramer of natural core streptavidin (S4) were measured at pH 7 and temperatures ranging from 15 to 45 °C using electrospray ionization mass spectrometry (ESI-MS). Two different approaches to data analysis were employed, one based on the initial rate of dissociation of the (S4 + 4B) complex, the other involving nonlinear fitting of the time-dependent relative abundances of the (S4 + iB) species. The two methods were found to yield koff values that are in good agreement, within a factor of two. The Arrhenius parameters for the dissociation of the biotin–streptavidin interaction in solution were established from the koff values determined by ESI-MS and compared with values measured using a radiolabeled biotin assay. Importantly, the dissociation activation energies determined by ESI-MS agree, within 1 kcal mol–1, with the reported value. In addition to providing a quantitative measure of koff, the results of the ESI-MS measurements revealed that the apparent cooperative distribution of (S4 + iB) species observed at short reaction times is of kinetic origin and that sequential binding of B to S4 occurs in a noncooperative fashion with the four ligand binding sites being kinetically and thermodynamically equivalent and independent.
Date created
2013
DOI
doi:10.7939/R3930P741
License information
© 2013 Deng, L., Kitova, E. N., & Klassen, J. S. This version of this article is open access and can be downloaded and shared. The original author(s) and source must be cited.
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Citation for previous publication
Deng, L., Kitova, E. N., & Klassen, J. S. (2013). Dissociation kinetics of the streptavidin-biotin interaction measured using direct electrospray ionization mass spectrometry analysis. Journal of the American Society for Mass Spectrometry, 24(1), 49-56.  http://doi.org/10.1007/s13361-012-0533-5

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File format: pdf (Portable Document Format)
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File title: Comparison of the Dissociation Kinetics of the Streptavidin-Biotin Interaction in Solution and in the Gas Phase
File author: John Klassen
Page count: 30
File language: en-US
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