Toxins have evolved to target regions of membrane ion channels that underlie ligand binding, gating, or ion permeation, and have thus served as invaluable tools for probing channel structure and function. Here, we describe a peptide toxin from the Earth Tiger tarantula that selectively and irreversibly activates the capsaicin- and heat-sensitive channel, TRPV1. This high-avidity interaction derives from a unique tandem repeat structure of the toxin that endows it with an antibody-like bivalency. The " double-knot" toxin traps TRPV1 in the open state by interacting with residues in the presumptive pore-forming region of the channel, highlighting the importance of conformational changes in the outer pore region of TRP channels during activation.
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We thank Ben Myers for cloning the frog TRPV1 cDNA and making it available for this study, and Roger Nicoll, Alex Chesler, Julio Cordero, Erhu Cao, and other members of our lab for helpful criticism and reading of the manuscript. We also thank the University of California, San Francisco, Mass Spectrometry Resource for instrumentation and technical assistance, supported by the National Institutes of Health (NIH) National Center for Research Resources. This work was supported by a NIH/National Institute of Neurological Disorders and Stroke (NINDS) Ruth Kirschstein predoctoral fellowship (C.B.), postdoctoral fellowships from the Damon Runyon Cancer Research Foundation (A.P.) and the International Human Frontier Science Program Organization (J.S.), and grants from the NIH/NINDS (D.J.).