ABSTRACT
Defined as a state function representing an inhibitor’s absolute affinity for its target enzyme, the experimentally determined enzyme inhibition constant (Ki) is widely used to rank order binding affinities of different inhibitors for a common enzyme or different enzymes for a common inhibitor and to benchmark computational approaches to predicting binding affinity. Herein, we report that adsorption of bis(7)-tacrine to the glass container surface increased its Ki against Electrophorus electricus acetylcholinesterase (eeAChE) to 3.2 ± 0.1 nM (n = 5) compared to 2.9 ± 0.4 pM (n = 5) that was determined using plastic containers with other assay conditions kept the same. We also report that, due to binding or “adsorption” of bis(7)-tacrine to the inactive eeAChE, the bis(7)-tacrine Ki increased from 2.9 ± 0.4 pM (n = 5) to 734 ± 70 pM (n = 5) as the specific eeAChE activity decreased from 342 U/mg to 26 U/mg while other assay conditions were kept the same. These results caution against using Kis to rank order binding potencies, define selectivity, or benchmark computational methods without knowing detailed assay conditions.
- Ki
- enzyme inhibition constant
- AChE
- acetylcholinesterase
- eeAChE
- Electrophorus electricus AChE
- ATCh
- acetylthiocholine chloride
- bis(7)-tacrine
- 1,7-N-heptylene-bis-9,9'-amino-1,2,3,4-tetrahydro-acridinium dihydrochloride
- DTNB
- 5,5’-dithiobis(2-nitrobenzoic acid)
- SEA
- specific enzyme activity
- tacrine
- 9-amino-1,2,3,4-tetrahydroacridinium monohydrochloride.