DATA AVAILABILITY STATEMENT
The data supporting the findings of this study are available from the
corresponding authors upon reasonable request. Some data may not be made
available because of privacy or ethical restrictions.
Background and Purpose: Local anesthetics block sodium and a
variety of potassium channels. Although previous studies identified a
residue in the pore signature sequence together with three residues in
the S6 segment as a putative binding site, the precise molecular basis
of Kv potassium channel inhibition by local anesthetics remained
unknown. Kv crystal structures predict that some of these residues point
away from the central cavity and face into a drug binding site called
‘side pockets´. Thus, the question arises whether the binding site of
local anesthetics is exclusively located in the central cavity or also
involves the ‘side pockets´.
Experimental Approach: A systematic functional alanine
mutagenesis approach, scanning 58 mutants, in concert with in
silico docking experiments and molecular dynamics simulations were
utilized to elucidate the binding site of bupivacaine and ropivacaine.
Key Results: Kv1.5 inhibition by local anesthetics requires
binding to the central cavity and the ‘side pockets´, where the latter
requires interactions with residues of the S5 and the backside of the S6
segment. Mutations in the ‘side pockets´ remove stereoselectivity of
Kv1.5 inhibition by bupivacaine. Strikingly, while we found that binding
to the ‘side pockets´ is conserved for the different local anesthetics,
the binding mode in the central cavity and the ‘side pockets´ shows
considerable variations.
Conclusion and Implications: Local anesthetics bind to the
central cavity and the ‘side pockets´ which provides a crucial key for
the molecular understanding of their Kv channel affinity and
stereoselectivity, as well as their spectrum of side effects.
Keywords: local anesthetics, bupivacaine, ropivacaine, Kv1
channels, side pockets, stereoselectivity