Analysis of Hormone-Protein Binding in Solution by Ultrafast Affinity Extraction

Interactions of Testosterone with Human Serum Albumin and Sex Hormone Binding Globulin

Xiwei Zheng, Cong Bi, Marissa Brooks, David S Hage

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Ultrafast affinity extraction was used to study hormone-protein interactions in solution, using testosterone and its transport proteins human serum albumin (HSA) and sex hormone binding globulin (SHBG) as models. Both single column and two-dimensional systems based on HSA microcolumns were utilized to measure the free fraction of testosterone in hormone/protein mixtures at equilibrium or that were allowed to dissociate for various lengths of time. These data were used to determine the association equilibrium constants (Ka) or global affinities (nKa′) and dissociation rate constants (kd) for testosterone with soluble HSA and SHBG. This method was also used to measure simultaneously the free fraction of testosterone and its equilibrium constants with both these proteins in physiological mixtures of these agents. The kd and Ka values obtained for HSA were 2.1-2.2 s-1 and 3.2-3.5 × 104 M-1 at pH 7.4 and 37°C. The corresponding constants for SHBG were 0.053-0.058 s-1 and 0.7-1.2 × 109 M-1. All of these results gave good agreement with literature values, indicating that this approach could provide information on a wide range of rate constants and binding strengths for hormone-protein interactions in solution and at clinically relevant concentrations. The same method could be extended to alternative hormone-protein systems or other solutes and binding agents.

Original languageEnglish (US)
Pages (from-to)11187-11194
Number of pages8
JournalAnalytical Chemistry
Volume87
Issue number22
DOIs
StatePublished - Oct 20 2015

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Sex Hormone-Binding Globulin
Serum Albumin
Testosterone
Hormones
Equilibrium constants
Proteins
Rate constants
Carrier Proteins
Protein Binding
Association reactions

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Analysis of Hormone-Protein Binding in Solution by Ultrafast Affinity Extraction : Interactions of Testosterone with Human Serum Albumin and Sex Hormone Binding Globulin. / Zheng, Xiwei; Bi, Cong; Brooks, Marissa; Hage, David S.

In: Analytical Chemistry, Vol. 87, No. 22, 20.10.2015, p. 11187-11194.

Research output: Contribution to journalArticle

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