Solubilisation and partial purification of auxin-binding sites of corn membranes

Abstract

THE demonstration by Hertel et al.¹ of reversible, high-affinity binding of auxins to membranous preparations from corn coleoptiles provided for the first time a reliable and readily accessible system for the study of potential receptor–phytohormone interactions. In a detailed investigation of the binding kinetics and specificity of this system, we presented evidence for the presence of two classes of auxin-binding sites with differing affinities and specificities^2,3. Site 1, with a K_D for the synthetic auxin 1-naphthalene-acetic acid (NAA) of 0.14 µM, was able to bind both auxins and physiologically inactive analogues, whereas site (K_D for NAA = 1.7 µM) displayed binding specificity compatible with that expected of an auxin receptor² and was located in a heavier membrane population, enriched in plasma membrane³. Auxin-binding activity can be readily solubilised from the membranes with Triton X-100 (ref. 2), but our attempts to purify such extracts further have met with only limited success. I report here that both binding sites can be conveniently removed from the membranes without detergent and purified approximately 100-fold in two steps.