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Abstract

Colicins are bacterial protein toxins produced by half of E. coli natural isolates that kill sensitive E. coli cells. Colicin M (Cma) inhibits incorporation of murein precursors into murein. Cma producer cells are protected by co-synthesis of an immunity protein, Cmi, that is located at the Cma target site in the periplasm and anchored to the cytoplasmic membrane by an N-terminal hydrophobic sequence [1]. We resumed our previous studies on Cma and Cmi after we had discovered that Cma activity requires the periplasmic FkpA prolyl cis-trans isomerase /chaperone [2]. Since the hydrophobic sequence is not essential for Cmi activity [1], crystallization was performed with a soluble Cmi that lacked the N- terminus. Cmi crystals were obtained under several conditions but only one single crystal diffracted to a resolution of 1.95 Å. By using the recently published software package ARCIMBOLDO [3], we succeeded to solve the structure by this de novo approach (Dayté Rodriguez, Isabel Usón- Finkenzeller, Instituto di Biología Molecular de Barcelona, Barcelona, Spain). In the crystal Cmi forms a dimer that is interlinked by a disulfide bridge. It is a highly charged protein with a surplus of negative charges presumably responsible for interaction with Cma which contains a cluster of positive charges.