PDBsum entry: 2cb6

Toxin

PDB id: 2cb6

Contents

Protein chains

(+ 4 more) 238 a.a.

(+ 0 more) 236 a.a.

PDB id:

2cb6

Name:

Toxin

Title:

Crystal structure of the catalytic domain of the mosquitocidal toxin from bacillus sphaericus, mutant e195q

Structure:

Mosquitocidal toxin. Chain: a, b, c, d, e, f, g, h, i, j, k, l, m, n, o, p. Fragment: catalytic domain residues 30-308. Engineered: yes. Mutation: yes

Source:

Bacillus sphaericus. Organism_taxid: 1421. Strain: ssii-1. Expressed in: escherichia coli. Expression_system_taxid: 511693.

Biol. unit:

Tetramer (from PDB file)

Resolution:

3.00Å R-factor: 0.227 R-free: 0.245

Authors:

D.J.Reinert,I.Carpusca,K.Aktories,G.E.Schulz

Key ref:

D.J.Reinert et al. (2006). Structure of the Mosquitocidal Toxin from Bacillus sphaericus. J Mol Biol, 357, 1226-1236. PubMed id: 16483607 DOI: 10.1016/j.jmb.2006.01.025

Date:

29-Dec-05 Release date: 22-Feb-06

Protein chains

Q03988  (Q03988_LYSSH) -  Mosquitocidal toxin

Seq: 870 a.a.

Struc: 238 a.a.*

Protein chains

Q03988  (Q03988_LYSSH) -  Mosquitocidal toxin

Seq: 870 a.a.

Struc: 236 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DOI no: 10.1016/j.jmb.2006.01.025

J Mol Biol 357:1226-1236 (2006)

PubMed id: 16483607

Structure of the Mosquitocidal Toxin from Bacillus sphaericus.

D.J.Reinert, I.Carpusca, K.Aktories, G.E.Schulz.

ABSTRACT

The catalytic domain of a mosquitocidal toxin prolonged by a C-terminal 44 residue linker connecting to four ricin B-like domains was crystallized. Three crystal structures were established at resolutions between 2.5A and 3.0A using multi-wavelength and single-wavelength anomalous X-ray diffraction as well as molecular replacement phasing techniques. The chainfold of the toxin fragment corresponds to those of ADP-ribosylating enzymes. At pH 4.3 the fragment is associated in a C(7)-symmetric heptamer in agreement with an aggregate of similar size observed by size-exclusion chromatography. In two distinct crystal forms, the heptamers formed nearly spherical, D(7)-symmetric tetradecamers. Another crystal form obtained at pH 6.3 contained a recurring C(2)-symmetric tetramer, which, however, was not stable in solution. On the basis of the common chainfold and NAD(+)-binding site of all ADP-ribosyl transferases, the NAD(+)-binding site of the toxin was assigned at a high confidence level. In all three crystal forms the NAD(+) site was occupied by part of the 44 residue linker, explaining the known inhibitory effect of this polypeptide region. The structure showed that the cleavage site for toxin activation is in a highly mobile loop that is exposed in the monomer. Since it contains the inhibitory linker as a crucial part of the association contact, the observed heptamer is inactive. Moreover, the heptamer cannot be activated by proteolysis because the activation loop is at the ring center and not accessible for proteases. Therefore the heptamer, or possibly the tetradecamer, seems to represent an inactive storage form of the toxin.

Selected figure(s)

Figure 1.
Figure 1. Stereoview of a ribbon representation of MTX30-308 from crystal form I, including the activation loop conformation (green) observed in crystal form II. The proper catalytic core of several bacterial toxins is marked in blue. The b-strands and a-helices are labeled, the C-terminal linker is red. The ARTT motif (pink) and the loop at position 150 (yellow) are likely to contact the target molecule. The positions of the two glutamate residues of the 195E-x-E motif are shown as balls. The labels N, AL and C mark residues 33, 264 and 298, respectively. The NAD^+ molecule was transferred from its binding location in cholera toxin26 to MTX30-308. In MTX30-308 its position is occupied by the inhibitory linker.

Figure 4.
Figure 4. Stereoview of the catalytic center of MTX30-308. (a) Detailed binding structure of the inhibitory linker. The crucial segment of the linker is 273D-F-D-M-F-A-P-N-G, which occupies the putative NAD^+ binding site. (b) Putative NAD^+-binding site as derived from cholera toxin.26 NAD^+ superimposes with the inhibitory linker explaining the competitive type of inhibition.17 The suggested interactions of NAD^+ with the protein (red hydrogen bonds) correspond to the common features of the ADP-ribosyl transferase family.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2006, 357, 1226-1236) copyright 2006.

Figures were selected by an automated process.