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(Strept)Avidin-Biotin: Two Interrelated Experiments for the Introductory Chemistry Laboratory

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David E. Hansen , Dengda Tang , Jon A. Sanborn and Mark D. Marshall

Department of Chemistry, Amherst College, Amherst, MA 01002

J. Chem. Educ., 2006, 83 (5), p 777

DOI: 10.1021/ed083p777

Publication Date (Web): May 1, 2006

Section:

History, Education, and Documentation

Abstract

A two-experiment sequence focusing on the noncovalent complex between the egg white protein avidin (or the similar protein streptavidin, which is expressed by the bacterium Streptomyces avidinii) and the essential cofactor biotin has been developed. The association equilibrium constant for the formation of the (strept)avidin–biotin complex is 1015, one of the largest ever measured for a noncovalent interaction between a protein and small molecule. In the first of the two experiments, students employ molecular modeling techniques to examine a series of noncovalent interactions, beginning with the argon dimer and culminating with (strept)avidin–biotin complex. In the second experiment, students titrate a solution of avidin with the dye 2-[(4-hydroxyphenyl)azo]benzoic acid (HABA). HABA has an absorption maximum at 348 nm, but upon binding to avidin the maximum redshifts to 500 nm, thus allowing the quantity of complex present to be determined. From these data, the equilibrium constant for the binding of HABA to avidin is calculated.

Keywords (Audience):

First-Year Undergraduate / General

Keywords (Domain):

Biochemistry

Keywords (Feature):

Molecular Modeling Exercises and Experiments

Keywords (Pedagogy):

Computer-Based Learning

Keywords (Subject):

Bioorganic Chemistry

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This article has been cited by 1 ACS Journal articles (1 most recent appear below).

Complementary Spectroscopic Assays for Investigating Protein−Ligand Binding Activity: A Project for the Advanced Chemistry Laboratory
David P. Mascotti and Mark J. Waner
Journal of Chemical Education2010 87 (7), 735-738
- Complementary Spectroscopic Assays for Investigating Protein−Ligand Binding Activity: A Project for the Advanced Chemistry Laboratory
  David P. Mascotti and Mark J. Waner
  Journal of Chemical Education2010 87 (7), 735-738
  A protein−ligand binding, guided-inquiry laboratory project with potential application across the advanced undergraduate curriculum is described. At the heart of the project are fluorescence and spectrophotometric assays utilizing biotin-4-fluorescein and ...
  Abstract | Full Text HTML | Hi-Res PDF | PDF w/ Links

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Received: August 03, 2009

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Lab-Expt

(Strept)Avidin-Biotin: Two Interrelated Experiments for the Introductory Chemistry Laboratory