Abstract
Voltage-gated potassium (Kv) channels enable potassium efflux and membrane repolarization in excitable tissues. Many Kv channels undergo a progressive loss of ion conductance in the presence of a prolonged voltage stimulus, termed slow inactivation, but the atomic determinants that regulate the kinetics of this process remain obscure. Using a combination of synthetic amino acid analogs and concatenated channel subunits we establish two H-bonds near the extracellular surface of the channel that endow Kv channels with a mechanism to time the entry into slow inactivation: an intra-subunit H-bond between Asp447 and Trp434 and an inter-subunit H-bond connecting Tyr445 to Thr439. Breaking of either interaction triggers slow inactivation by means of a local disruption in the selectivity filter, while severing the Tyr445-Thr439 H-bond is likely to communicate this conformational change to the adjacent subunit(s). DOI: http://dx.doi.org/10.7554/eLife.01289.001.
Highlights
Enzymes and catalytic proteins have evolved to balance the thermodynamic challenges of stability and substrate throughput (Shoichet et al, 1995)
One previously proposed (Cordero-Morales et al, 2011; Hoshi and Armstrong, 2013), but untested possibility is that a H-bond is formed between the hydrogen on the indole nitrogen of Trp434 and the carboxylate moiety of Asp447, and disrupting this H-bond would promote conformational changes associated with slow inactivation
While structural data suggests a possible H-bond between Trp434 and Asp447, the available functional data cannot definitely discriminate between the roles of side chain size and/or volume or hydrogen bonding ability being the major determinant of slow inactivation at position 434
Summary
Enzymes and catalytic proteins have evolved to balance the thermodynamic challenges of stability and substrate throughput (Shoichet et al, 1995). Inactivation can be described by two kinetically and mechanistically distinct processes termed fast (or ‘N-type’) inactivation and slow (or ‘C-type’) inactivation (Hoshi et al, 1991; Kurata and Fedida, 2006; Hoshi and Armstrong, 2013) While the former results from a channel peptide docking within the open cytoplasmic entrance to the permeation pathway (Hoshi et al, 1990, 1991; Zhou et al, 2001), the latter is assumed to involve highly cooperative local conformational changes near the selectivity filter, a notion supported by electrophysiological, structural and computational approaches
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