PDBsum entry 1v74

Antibiotic/immune system

PDB id: 1v74

Contents

Protein chains

107 a.a.

87 a.a.

Ligands

1PE

Waters ×195

References listed in PDB file

Key reference

Title

Structural inhibition of the colicin d trnase by the trna-Mimicking immunity protein.

Authors

M.Graille, L.Mora, R.H.Buckingham, H.Van tilbeurgh, M.De zamaroczy.

Ref.

EMBO J, 2004, 23, 1474-1482. [DOI no: 10.1038/sj.emboj.7600162]

PubMed id

15014439

Abstract

Colicins are toxins secreted by Escherichia coli in order to kill their competitors. Colicin D is a 75 kDa protein that consists of a translocation domain, a receptor-binding domain and a cytotoxic domain, which specifically cleaves the anticodon loop of all four tRNA(Arg) isoacceptors, thereby inactivating protein synthesis and leading to cell death. Here we report the 2.0 A resolution crystal structure of the complex between the toxic domain and its immunity protein ImmD. Neither component shows structural homology to known RNases or their inhibitors. In contrast to other characterized colicin nuclease-Imm complexes, the colicin D active site pocket is completely blocked by ImmD, which, by bringing a negatively charged cluster in opposition to a positively charged cluster on the surface of colicin D, appears to mimic the tRNA substrate backbone. Site-directed mutations affecting either the catalytic domain or the ImmD protein have led to the identification of the residues vital for catalytic activity and for the tight colicin D/ImmD interaction that inhibits colicin D toxicity and tRNase catalytic activity.

Figure 1.
Figure 1 Stereo views. (A) Ribbon representation of the complex between colicin D (yellow) and ImmD (pink). The colicin D His611 and the ImmD Glu56 side chains at the interface are shown as sticks. Residues whose mutation has no effect on the in vivo and in vitro activities are highlighted in green. (B) Close-up of the colicin D putative active site. The colicin D tRNase domain is shown as a ribbon with the potential active site residues represented as sticks. For clarity, residues are identified by their one-letter code in all figures. (C) Close-up of main colicin D (yellow) and ImmD (green) residues involved at the dimer interface.

Figure 2.
Figure 2 Molecular surface representation. (A, B) Secondary structure assignments along the colicin D (A) and ImmD (B) sequences. Residues involved in complex formation are highlighted in bold. (C) The binary colicin D (yellow) -ImmD (green) complex. The orientation is the same as in Figure 1A. The red-coloured region corresponds to ImmD positions located outside the interface, but whose mutation abolishes or reduces the inhibitory capacity. (D) Surface mapping of the colicin D catalytic active site residues. Unmutated positions are coloured grey. Residues whose mutation does not alter tRNase activity are shown in green, those whose substitution results in partially or fully inactivated proteins are depicted in orange and red, respectively. For clarity, only residues whose mutation results in complete loss of activity are labelled. (E) ImmD foot-printing (green) at the colicin D surface. The His611 residue is coloured yellow. The orientation is the same as in (D). (F) Open leaflet representation of the complex illustrating the electrostatic complementarity between colicin D and ImmD. Regions of the surface with negative potential are coloured red and those with positive potential are in blue. The colicin D orientation is similar as in (D). The ImmD is rotated 180° around the vertical axis. (G) Surface mapping of the ImmD residues involved in the inhibitory effect. Colour coding is the same as in (D). The orientations of the ImmD in the top and bottom panels are related by a 180° rotation around the vertical axis.

The above figures are reprinted from an Open Access publication published by Macmillan Publishers Ltd: EMBO J (2004, 23, 1474-1482) copyright 2004.