Publication: An experiment illustrating the change in ligand pK<inf>a</inf>upon protein binding
Issued Date
2012-05-08
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00219584
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2-s2.0-84860746126
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Mahidol University
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SCOPUS
Bibliographic Citation
Journal of Chemical Education. Vol.89, No.6 (2012), 791-795
Suggested Citation
Pirom Chenprakhon, Bhinyo Panijpan, Pimchai Chaiyen An experiment illustrating the change in ligand pK<inf>a</inf>upon protein binding. Journal of Chemical Education. Vol.89, No.6 (2012), 791-795. doi:10.1021/ed2006482 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/13969
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An experiment illustrating the change in ligand pK<inf>a</inf>upon protein binding
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Abstract
The modulation of ligand pK a due to its surrounding environment is a crucial feature that controls many biological phenomena. For example, the shift in the pK a of substrates or catalytic residues at enzyme active sites upon substrate binding often triggers and controls enzymatic reactions. In this work, we developed an experiment using spectrophotometric method to demonstrate how ligand pK a values can be influenced by specific interactions in the protein-binding pocket using riboflavin binding protein (RP) and its ligands (riboflavin, RF, and neutral red, NR). A direct plot of observed absorbance versus pH was analyzed by nonlinear regression. The pK a values of free and RP-bound RF were determined to be 10.0 ± 0.1 and ∼13.3, respectively, and the pK a values of free and RP-bound NR were 6.8 ± 0.1 and 7.8 ± 0.1, respectively. This laboratory clearly demonstrates that the environment of a protein-binding site can affect the pK a value of a ligand. The experiment can be adapted or used as-is for undergraduate students in biochemistry or chemistry (analytical or physical chemistry) or first-year graduate students in biochemistry and related fields. © 2012 The American Chemical Society and Division of Chemical Education, Inc.