Synthetic Analogues of the Snail Toxin 6-Bromo-2-mercaptotryptamine Dimer (BrMT) Reveal That Lipid Bilayer Perturbation Does Not Underlie Its Modulation of Voltage-Gated Potassium Channels

Abstract

Drugs do not act solely by canonical ligand-receptor binding interactions. Amphiphilic drugs partition into membranes thereby perturbing bulk lipid bilayer properties and possibly altering the function of membrane proteins. Distinguishing membrane perturbation from more direct protein-ligand interactions is an ongoing challenge in chemical biology. Herein, we present one strategy for doing so, using dimeric 6-bromo-2-mercaptotryptamine (BrMT) and synthetic analogs. BrMT is a chemically unstable marine snail toxin that has unique effects on voltage-gated K+ channel proteins, making it an attractive medicinal chemistry lead. BrMT is amphiphilic and perturbs lipid bilayers, raising the question of whether its action against K+ channels is merely a manifestation of membrane perturbation. To determine whether medicinal chemistry approaches to improve BrMT might be viable, we synthesized BrMT and 11 analogs and determined their activities in parallel assays measuring K+ channel activity and lipid bilayer properties. Structure-activity relationships were determined for modulation of the Kv1.4 channel, bilayer partitioning, and bilayer perturbation. Neither membrane partitioning, nor bilayer perturbation, correlate with K+ channel modulation. We conclude that BrMT’s membrane interactions are not critical for its inhibition of Kv1.4 activation. Further, we found that alkyl or ether linkages can replace the chemically labile disulfide bond in the BrMT pharmacophore, and we identified additional regions of the scaffold that are amenable to chemical modification. Our work demonstrates a strategy for determining if drugs act by specific interactions or bilayer-dependent mechanisms, and chemically stable modulators of Kv1 channels are reported.

Antidepressants are modifiers of lipid bilayer properties

“…That is, though the gating mechanisms of channels formed by integral membrane proteins and gA channels are different, they are sensitive to the same changes in bilayer properties. In cases where there is no correlation between the effects in cells and in the gA-based assays (e.g., Herold et al, 2017; Dockendorff et al, 2018), the parsimonious interpretation becomes that the compounds alter membrane protein function by direct interactions or binding to their target.…”

“…Notably, TCAs and SSRIs have low-affinity interactions with a diverse group of membrane proteins (Rammes and Rupprecht, 2007; Bianchi, 2008; see also Table S1), which may provide additional mechanisms for altering neural chemistry and circuitry (Duman et al, 2016). The mechanisms underlying this polypharmacology remain unclear, but integral membrane proteins have two things in common: they are membrane spanning, and their activity can be modulated by changes in lipid bilayer properties (curvature, thickness, and elasticity) that can be induced by the addition of amphiphiles, including many biologically active molecules (e.g., a variety of toxins; Suchyna et al, 2004; Dockendorff et al, 2018), currently used or discontinued drugs (Rusinova et al, 2011, 2015), and phytochemicals (Ingólfsson et al, 2014; see also Lundbæk et al, 2010b, Table 3).…”

Conformational dynamics between transmembrane domains and allosteric modulation of a metabotropic glutamate receptor

“…Finally, in contrast to BINA, the NAM Ro 64 showed fast ON and slow OFF kinetics despite the lack of membrane perturbation in an in vitro liposome assay. It’s worth noting that while one would expect that very hydrophobic molecules, such as Ro 64, would be potent bilayer modifiers that is not always the case (Alejo et al, 2013; Dockendorff et al, 2018).…”

hetN and patS Mutations Enhance Accumulation of Fatty Alcohols in the hglT Mutants of Anabaena sp. PCC 7120

“…In some marine bacteria, PKS synthesize very long chain fatty acids, such as docosahexaenoic acids ( Okuyama et al, 2007 ). Recently, NRPSs have successfully been expressed in Anabaena for bioproduction ( Videau et al, 2016 , 2019 ). The replacement of genes involved in the synthesis of Hgl with desired PKS/NRPS will be of great interest.…”