W0273

Structure of the E. coli Outer Membrane Receptor, BtuB, and the Receptor Binding Domain of Colicin E3. W. A. Cramer, M. Zhalnina, S. Bano^†, S. D. Zakharov, M Wiener¹, & G. Kurisu^‡,Dept. of Biological Sciences, Purdue Univ., West Lafayette, IN 47907, ¹Dept. of Molecular Physiology and Biological Physics, Univ. of Virginia, Charlottesville, VA 22908, USA, permanent address: ^†HEJ Research Inst. of Chemistry, Univ. of Karachi, Karachi, Pakistan; ^‡Inst. for Protein Research, Osaka Univ., Suita, Osaka 565-0871, Japan.

The structure of a nM affinity complex between the coiled-Alacoil 135 residue receptor binding domain of colicin E3 (Soelaiman et al., 2001) and the colicin-vitamin B₁₂ receptor has been solved to 2.75 Å [Space group P2₁2₁2₁, a = 76.93, b = 80.10 and c = 233.60 Å; solvent content (1 BtuB-R135 complex in asymmetric unit, 71.4 % (V_M = 4.3 Å³Da^-1); R_cryst = 0.234; R_free = 0.283) by molecular replacement method using coordinates of apo-BtuB (Chimento et al., 2003). Bound R135, involving the receptor-proximal 31 residues of R135 and the extracellular loop region of BtuB, extends obliquely from BtuB, at an angle of 40° from the membrane plane. Residue M383 of R135 penetrates to the top of the BtuB “cork” region. Structure changes induced by binding, in which interactions of R135 residues R399 and W390, respectively, with BtuB-E330 and -W405 are prominent, cause loops N228-D241 and Y277-A288 of BtuB to reorder, and 10 residues at each terminus of the R135 coiled-coil, to reorder. Superimposition of the whole colicin E3 structure with the R135 region of the complex shows that the translocation domain of the colicin would be close to the membrane, or a second receptor. This structure, and the absence of a large channel conductance for BtuB, imply that BtuB serves as a receptor for the colicin, but does not translocate the colicin across the OM.