Publication: An experiment illustrating the change in ligand pKa upon protein binding
8
Issued Date
2012-02-28
Resource Type
Language
eng
Rights
Mahidol University
Rights Holder(s)
American Chemical Society and Division of Chemical Education, Inc.
Bibliographic Citation
Journal of Chemical Education. Vol.89, No.6 (2012), 791-795
Suggested Citation
Pirom Chenprakhon, ภิรมย์ เชนประโคน, Bhinyo Panijpan, Pimchai Chaiyen An experiment illustrating the change in ligand pKa 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/123456789/3391
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Title
An experiment illustrating the change in ligand pKa upon protein binding
Abstract
The modulation of ligand pKa
due to its surrounding
environment is a crucial feature that controls many biological phenomena.
For example, the shift in the pKa
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 pKa
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 pKa
values of free and RP-bound RF were
determined to be 10.0 ± 0.1 and
∼
13.3, respectively, and the pKa
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 pKa
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.
Sponsorship
Institute for the Promotion of
Teaching Science and Technology (IPST), Thailand (to P.
Chenprakhon), and grants from the Thailand Research Fund
(BRG5480001) and from the Faculty of Science, Mahidol
University (to P. Chaiyen)