Evolution of metal selectivity in templated protein interfaces

Abstract

Selective binding by metalloproteins to their cognate metal Ions is essential to cellular survival. How proteins originally acquired the ability to selectively bind metals and evolved a diverse array of metalcentered functions despite the availability of only a few metal-coordinating functionalities remains an open question. Using a rational design approach (Metal-Templated Interface Redesign), we describe the transformation of a monomeric electron transfer protein, cytochrome cb562, into a tetrameric assembly ( C96RIDC-14) that stably and selectively binds Zn 2+ and displays a metal-dependent conformational change reminiscent of a signaling protein. A thorough analysis of the metal binding properties ofC96RIDC-14 reveals that It can also stably harbor other divalent metals with affinities that rival (Ni2+) or even exceed (Cu2+) those of Zn2+ on a per site basis. Nevertheless, this analysis suggests that our templating strategy simultaneously introduces an increased bias toward binding a higher number of Zn2+ ions (four high affinity sites) versus Cu2+ or Ni2+ (two high affinity sites), ultimately leading to the exclusive selectivity of C96RIDC14 for Zn2+ over those ions. More generally, our results indicate that an initial metal-driven nucleation event followed by the formation of a stable protein architecture around the metal provides a straightforward path for generating structural and functional diversity. © 2010 American Chemical Society.