Allostery is a fundamental principle of protein regulation that remains hard to engineer, particularly in membrane proteins such as ion channels. Here we use human Inward Rectifier K + Channel Kir2.1 to map site-specific permissibility to the insertion of domains with different biophysical properties. We find that permissibility is best explained by dynamic protein properties, such as conformational flexibility. Several regions in Kir2.1 that are equivalent to those regulated in homologs, such as G-protein-gated inward rectifier K + channels (GIRK), have differential permissibility; that is, for these sites permissibility depends on the structural properties of the inserted domain. Our data and the well-established link between protein dynamics and allostery led us to propose that differential permissibility is a metric of latent allosteric capacity in Kir2.1. In support of this notion, inserting light-switchable domains into sites with predicted latent allosteric capacity renders Kir2.1 activity sensitive to light.
Bibliographical noteFunding Information:
We thank Matthew R. Whorton, Mikael Elias, Sivaraj Sivaramakrishnan, and the entire Schmidt Lab for helpful feedback and discussion, Therese Martin with flow cytometry technical advice and support, David Savage and Avi Flamholz for technical advice and development of the DIPseq alignment pipeline, Alina Zdechlik, Tejas Gupte, and Daniel Sorenson for helpful feedback on the paper, and Steffan Okorafor for assistance with single-mutant construction. W.C.-M. is funded by a National Science Foundation Graduate Research Fellowship.
© 2019, The Author(s).