The DNase domain of colicin E9 (E9 DNase) represents the 134 C-terminal-most residues of
intact colicin E9, with a methionine residue substituted for a lysine at the N-terminus.
In this study, isotopically labelled E9 DNase is in complex with its unlabelled
inhibitor protein, Im9. Thus, only the E9 DNase component is observed.

The crystal structure of the E9 DNase-Im9
complex contains a Ni2+ ion in the putative
active site of E9 DNase. In this study,
E9 DNase in complex with Im9 is metal-free
in solution.

Inhibitor (Im9)-bound DNase domain of colicin E9

The E9 DNase forms the cytotoxic domain of colicin E9 and is located at its C-terminus (the last 134 residues). It acts as an endonuclease and is active on both single- and double-stranded DNA but with undefined specificity. Colicins are plasmid-encoded protein antibiotics produced by, and active against, Eschericia coli and closely related bacteria. The cytotoxic activity of colicin E9 is neutralised by the binding of its 9.5 kDa cognate immunity protein, Im9, to the E9 DNase.

Assignments refer to the major conformational species of E9 DNase when bound to its inhibitor, Im9.

Assignments refer to the minor conformational species of E9 DNase when bound to its inhibitor, Im9.

Whittaker, S. B.-M., Czisch, M., Wechselberger, R., Kaptein, R., Hemmings, A. M., James, R., Kleanthous, C., and Moore, G. R., "Slow Conformational Dynamics of an Endonuclease Persist in its Complex with its Natural Protein Inhibitor," Protein Sci. 9, 713-720 (2000).
Slow Conformational Dynamics of an Endonuclease Persist in its Complex with its Natural Protein Inhibitor

The NMRPipe system is a UNIX software environment of processing, graphics, and analysis tools designed to meet current routine and research-oriented multidimensional processing requirements, and to anticipate and accommodate future demands and developments. The system is based on UNIX pipes, which allow programs running simultaneously to exchange streams of data under user control. In an NMRPipe processing scheme, a stream of spectral data flows through a pipeline of processing programs, each of which performs one component of the overall scheme, such as Fourier transformation or linear prediction. Complete multidimensional processing schemes are constructed as simple UNIX shell scripts. The processing modules themselves maintain and exploit accurate records of data sizes, detection modes, and calibration information in all dimensions, so that schemes can be constructed without the need to explicitly define or anticipate data sizes or storage details of real and imaginary channels during processing. The asynchronous pipeline scheme provides other substantial advantages, including high flexibility, favorable processing speeds, choice of both all-in-memory and disk-bound processing, easy adaptation to different data formats, simpler software development and maintenance, and the ability to distribute processing tasks on multi-CPU computers and computer networks.

Delaglio, F., Grzesiek, S., Vuister, G.W., Zhu, G., Pfeifer, J. & Bax, A. (1995)
J. Biomol. NMR 6, 277-293

NMRPipe: a multidimensional spectral processing system based on UNIX pipes.

Bartels, C., Xia, T.-H., Billeter, M., Guentert, P. & Wuethrich, K. (1995)
J. Biomol. NMR 6, 1-10

Eaton, T. & James, R. (1989) Nucleic Acids Res. 17, 1761-1761
Complete nucleotide sequence of the colicin E9 (cei) gene.

We have determined the nucleotide sequence of a 1500 bp fragment of the ColE9-J plasmid which encodes colicin E9 immunity and colicin E5 immunity and contains two lys genes. Open reading frames corresponding to the four genes have been located and their position confirmed by transposon mutagenesis of sub-clones of the ColE9-J plasmid. The E9imm gene shows 69% homology at both the nucleotide and the amino acid level to the previously sequenced E2imm gene. The E5imm gene shows little homology to any other E colicin immunity gene which has been sequenced. The lys gene distal to the 3' end of the E5imm gene shows considerable sequence homology to all other previously sequenced E colicin lys genes. The lys gene distal to the 3' end of the E9imm gene is identical to the pColE2 and pColE3 lys genes for the first 59 nucleotides but encodes a much smaller gene product than any other lys gene which has been sequenced. The two lys genes sequenced here are exceptions to Shepherd's rule concerning the number of RNY codons in the three possible reading frames.
James, R., Jarvis, M. & Barker, D.F. (1987) J. Gen. Microbiol. 133, 1553-1562.
Nucleotide sequence of the immunity and lysis region of the ColE9-J plasmid.

E9 DNase exists as two conformers in
solution (60:40) that undergo slow chemical
exchange. This conformational heterogeneity
remains upon inhibitor-binding.

Crystal Structure Of The Escherichia Coli Colicin E9 Dnase Domain With Its Cognate Immunity Protein Im9

Crystal Structure Of Colicin E9 Dnase Domain With Its Cognate Immunity Protein Im9 (1.7 Angstroms)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9(E41a)

Crystal Structure Of The E9 Dnase Domain

Crystal Structure Of The Mutant His103ala Of The Colicin E9 Dnase Domain In Complex With Zn+2 (2.0 Angstroms)

Crystal Structure Of The Colicin E9, Mutant His103ala, In Complex With Mg+2 And Dsdna (Resolution 2.9a)

Crystal Structure Of The Colicin E9, Mutant His103ala, In Complex With Zn+2 And Dsdna (Resolution 2.4a)

Crystal Structure Of The Complex Of The Colicin E9 Dnase Domain With A Mutant Immunity Protein, Imme9 (D51a)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9 (Y55a)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9 (Y54f)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9 (Y55f)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9 (V34a)

Crystal Structure Of The E9 Dnase Domain With A Mutant Immunity Protein Im9 (D51a)

Chemical Shift Structure Of Colicin E9 Dnase Domain With Its Cognate Immunity Protein Im9

N75a Mutant Of E9 Dnase Domain In Complex With Im9

K97a Mutant Of E9 Dnase Domain In Complex With Im9

R54a Mutant Of E9 Dnase Domain In Complex With Im9

F86a Mutant Of E9 Dnase Domain In Complex With Im9

The Crystal Structure Of Im2 In Complex With Colicin E9 Dnase

colicin E9 [Escherichia coli str. K-12 substr. W3110]

colcinin E9 C-terminal fragment [Plasmid ColE9-J]

To produce Im9-bound E9 DNase, purified unlabelled Im9 was added to uniformly 13C/15N-labelled E9 DNase.

Backbone 1H, 13C, and 15N Chemical Shift Assignments for the Inhibitor-Bound DNase Domain of Colicin E9.

homologous Immunity protein Im9 bound to DNase domain of colicin E9

DNase domain of non cognate binding partner E9

Simultaneous analysis of triple resonance spectra. Very useful for analysing multiple data sets simultaneously.

Defining strips for 3D NOESY to aid sequential assignment.