Toward a Molecular Lego Kit for Engineering Specialized Channels

“What I cannot create, I do not understand.” — Richard Feynman (1988)

An organism’s ability to sense the world ultimately relies on specialized proteins in its sensory neurons to probe the external world on behalf of the entire organism. Channels, a group of proteins that act as gatekeepers of ions, are often delegated to the front end of the job. As a result, highly specialized channels, such as those that sense odors, temperature, and even touch, have evolved in all corners of the world. Over the years, the genetic identities of many such channels have been demystified. Our current challenge lies in pinpointing the nanoscopic means by which they sense the world. To achieving this goal, an inevitable path is to locate the intramolecular modules (often referred to as domains) that grant channels their special ability to sense the environment. Several remarkable studies in recent years have made significant progress in attacking this problem.

KcsA

Only a few years ago, in 2011, the Sternson group exploited the properties of specialized domains to engineer new ligand-gated channels, which they called PSAMs2. First, the Sternson group made the critical observation that ligand-gated (i.e. molecule-sensing) ion channels can be divided into two somewhat independent domains, the ligand-binding domain and the ion channel domain. By screening candidate mutations in the ligand-binding domain of a starter channel, they were able to engineer the channel to lose its innate affinity to its natural ligand and acquire a preference for a synthetic molecule. By transplanting this new ligand binding domain onto other excitatory and inhibitory ion channel domains, the Sternson group successfully created novel excitatory and inhibitory channels. These channels now specialize in binding synthetic ligands that have never occurred in any biological system and are used as a tool to manipulate neuron activities.

Recent work by researchers in the Jan labs identified another elegant example of specialized domains in a touch-sensitive channel, NompC3. NompC has a long tail of short, repeated sequences known as Ankyrin repeats. These Ankyrin repeats connect the NompC channel, which resides on the surface of the cell, to the cell’s cytoskeleton, much as a ship’s anchor secures its vessel to the bottom of the ocean. When the cell surface is deformed by touch, it changes the distance between NompCs and the cytoskeleton, causing these Ankyrin chains to pull on the channels, just as a ship’s anchor will pull on the ship when ocean waves begin pushing the ship away. In the case of NompC, however, the Ankyrin chain can actually pull open the channel and, quite unexpectedly, plays an important role in defining the distance between the cell surface and cytoskeleton (i.e. the depth of the ocean in the ship analogy). Finally, transplanting the Ankyrin chain to a touch-insensitive channel renders the new channel touch-sensitive, just like the original NompC channel. The Ankyrin chain can therefore serve as a Lego piece for making touch-sensing channels.

Although specialized domains are common, they are not a requirement for specialized channels. For example, the search for a heat-sensitive domain in the thermosensitive TrpV1 channel has yielded largely non-overlapping regions of the protein. Facing this paradox, the Chanda group4 built on theoretical work that proposed that the temperature-gating properties of a channel might result from its general interaction with the cell membrane5. By changing the membrane-interaction properties of a small number of amino acids in a potassium channel, which is normally temperature insensitive, the researchers were able to create new channels that were just as temperature-sensitive as the natural ones. These manipulations also removed the potassium channel’s intrinsic voltage sensitivity, implying a shared mechanism between these two intuitively different senses.

Perhaps to Mr. Feynman’s disappointment, creating channels on a blackboard using only a handful of principles is still a dream of the future. In all three of these examples, rationality dictates the general directions of the research paths, but the details are left to hard work and serendipity. Nonetheless, the ability to grant new sensing properties to old channels by means of rational design should confer a great sense of achievement, as deserved by those who steal secrets from nature.

References:

  1. Doyle, D. A. et al. The structure of the potassium channel: molecular basis of K+ conduction and selectivity. Science 280, 69–77 (1998).
  1. Magnus, C. J. et al. Chemical and genetic engineering of selective ion channel-ligand interactions. Science 333, 1292–6 (2011).
  1. Zhang, W. et al. Ankyrin Repeats Convey Force to Gate the NOMPC Mechanotransduction Channel. Cell 162, 1391–1403 (2015).
  1. Chowdhury, S., Jarecki, B. W. & Chanda, B. A molecular framework for temperature-dependent gating of ion channels. Cell 158, 1148–58 (2014).
  1. Clapham, D. E. & Miller, C. A thermodynamic framework for understanding temperature sensing by transient receptor potential (TRP) channels. Proc. Natl. Acad. Sci. U. S. A. 108, 19492–7 (2011).